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					Annual Report
                                                    2002/03

Applied research and development at the interface
of life sciences and material sciences
                                                   NMI Natural and Medical Sciences
                                                   Institute at the University of Tübingen




Applied research and development at the
interface of life sciences and material sciences
NMI Overview
                                                 Table of contents




 2    NMI overview                                               38   Surface and interface technology
 3    Foreword                                                   40   Microsystems and nanotechnology
 4    Advisory board and executive management                    40   Sensor arrays and multiwells with microelectrodes
 5    Areas of core competence                                   41   Bio MEMS
 6    Good news                                                  42   Nano-Workbench
 7    Steering instruments                                       43   Focused ion beam (FIB)
 8    Network
 9    Start-up companies                                         44   Functional surfaces and layers
10    Patents                                                    44   Analysis of interfaces and microstructures
11    Public relations                                           45   Layers
12    Meeting clients expectations
13    Testing medical products according to                      47   Bonding, adhesion and tribological systems
      DIN EN ISO/IEC 17025                                       47   Bonding systems
                                                                 48   Tribological systems

14    Pharma and biotechnology
16    Functional genomics                                        49   Staff
16    Gene transfer                                              50   Publications
18    Protein-BioChips                                           51   Projects
20    A new Cell-BioChip                                         52   Roadmap to the NMI

21    Test systems for drug discovery
22    Electrophysiology with cells and tissues
23    Characterization of ion channels
25    Blood-brain-barrier

26    Bioanalytics and synthesis
26    Bioanalytics
27    Peptide synthesis



28    Biomedical technology
30    Neurotechnology
31    Retina implants
32    Neuro implants

33    Tissue engineering
33    Regeneration of joint cartilage
35    Regeneration of nerves

36    Development of medical products
37    Surface coating
37    Qualification of cleanliness




2    NMI Annual Report 2002/03
Foreword




The year 2003 was the fourth con-            We also like to thank the members                    mental impact on tax revenues, resulting
secutive year of growth for the NMI, an      of our advisory board for their advice               in decreased public funding for research.
impressive result in view of the difficult   and support, especially Prof. Hanack,                All these factors will require from us an
macro-economic environment. With             University of Tübingen, and Ministerial-             enormous effort in order to continue on
approximately 7 Mio Euro revenues            direktor Dr. Epple, Ministry of Economic             our path of sustainable growth.
and 90 employees, the institute has          Affairs, who have served as Chairman                 We are confident that also in the future,
now attained considerable national and       of the board.                                        through an efficient transfer of knowl-
international reputation. This growth        An important factor leading to our suc-              edge and technology from the NMI
is derived mainly from R&D and service       cess is the strong interaction with scien-           to industry, we will be able to support
contracts from industry, amounting to        tists and clinicians of the University of            our clients in their efforts to develop
3 Mio Euro in the year 2003. In particu-     Tübingen. Equally important is the inter-            innovative products, thus increasing their
lar the small and medium enterprises         action with members of the Science and               competitiveness and allowing the crea-
have turned to the NMI to develop new        Technology Park Tübingen-Reutlingen.                 tion of new jobs.
products. Our clients reside in Germany,
Europe and overseas. This reflects our       In the coming years we will face new
visibility and competence in various         and challenging tasks. With over 90
areas, such as biotechnology and medi-       employees the NMI has grown to a size
cal technology. We wish to thank our         which requires new and more adequate
customers for their confidence in the        structures and steering instruments. The             Enzio Müller
quality of our services.                     local biotechnology scene is still facing            Managing director
                                             difficult times. One of our preeminent
Notwithstanding the considerable public      goals will continue to be to support
budget constraints, the Ministry of          these biotechnology companies through
Economic Affairs of Baden-Württem-           our R&D competence. Furthermore, the
berg was able to support the NMI with        continuing consolidation in the phar-
approximately 1.5 Mio Euro. Unfortu-         maceutical industry will lead to struc-
nately this represents a further reduction   tural changes to which we will have to               Hugo Hämmerle
of public funding to 24% of our total        respond. The area of medical products                Deputy managing director
revenues. This makes it more and more        and medical technology is facing the
difficult to maintain and modernize our      pressures of cost-reduction measures in
technical infrastructure and to carry out    public health. The stagnant economy
the exploratory R&D which will later         in Germany is continuing to have a detri-
benefit our industrial clients, in par-
ticular small and medium enterprises in
our region. We are trying to buffer this
decreased support by the local govern-
ment by implementing those manage-
rial, operational and business measures
which are known to allow optimal
utilization of resources, thereby increas-
ing our efficiency and productivity.

Great recognition goes to our employ-
ees for their outstanding work. Their
engagement and dedication has led to a
high degree of customer satisfaction, to
innovation leading to numerous patents
and to our increasing international repu-
tation and visibility.                                          Enzio Müller, Managing Director           Hugo Hämmerle,
                                                                                                          Deputy managing director




                                                                                                                  NMI Annual Report 2002/03   3
NMI Overview
                                Advisory board and executive management




                                Executive body of the NMI



                                The NMI – Natural and Medical Sciences              Advisory Board
                                Institute – is a foundation (non-profit             Dr. K. Epple, chairman, Ministerial-
                                organization) established in 1985.                  direktor, Wirtschaftsministerium
                                The mission of the NMI is to transfer               Baden-Württemberg
                                the results of basic research into the              Prof. Dr. H. Brunner, Fraunhofer IGB,
                                economy. The president of the founda-               Stuttgart
                                tion and the managing director of the               Frau S. Dingfelder, Ministerium
                                Institute is Dr. Enzio Müller. An advisory          für Wissenschaft und Kunst
                                board supervises the activities of the              Baden-Württemberg
                                managing director.                                  Dr. W. Epp, IHK, Reutlingen
                                The NMI is financially and administra-              Dr. S. Hock, ZF Friedrichshafen AG
                                tively completely independent from                  Prof. Dr. K.-P. Jäckel, BASF AG,
                                the University of Tübingen. At the                  Ludwigshafen
                                operational level, however, there is                Prof. Dr. Dr. h. c. J. Löhn, Steinbeis-
                                a very intense collaboration. Numer-                Stiftung für Wirtschaftsförderung
                                ous research projects are carried out in            Baden-Württemberg
                                cooperation with scientists and clinicians          Prof. Dr. G. Obieglo, Rektor der
                                of the University. Graduate and under-              FH Reutlingen
                                graduate students perform their thesis              Frau B. Bosch, Oberbürgermeisterin
                                work at the NMI.                                    T. Reumann, Erster Bürgermeister,
                                                                                    Stadt Reutlingen
                                President and Vice-presidents of                    Prof. Dr. Dr. h. c. E. Schaich, Rektor
                                the foundation                                      Prof. Dr. T. Risler, Prorektor,
                                E. Müller, Ph.D., President                         Universität Tübingen
                                Dr. H. Hämmerle, Vice-president                     Prof. Dr. H. P. Rodemann,
                                Prof. Dr. D. Kern, Vice-president.                  Universität Tübingen
                                                                                    Dr. A. Siegel, Carl Zeiss AG,
                                                                                    Oberkochen
                                                                                    Dr. H. Stallforth, Aesculap AG &
                                                                                    Co KG, Tuttlingen
                                                                                    Dr. J. Traber, Bayer AG, Leverkusen
                                                                                    Prof. Dr. H. Weber,
                                                                                    Universität Tübingen
                                                                                    Prof. Dr. E. Zrenner,
                                                                                    Universität Tübingen




                                                      From the left:
                                                      R. Roscher (secretary), R. Rubow (Chief financial
                                                      officer), E. Müller (Managing director), I. Digel (secre-
                                                      tary) and H. Hämmerle (Deputy managing director).




4   NMI Annual Report 2002/03
Core-competence areas




Our 3 core-competence areas



The NMI is focused on pharma and bio-        Center for Biomaterials and Organ-
technology, biomedical technology and        Substitution (BMOZ), as well as the
surface and interface technology. The        Competence Center for Minimal Invasive
latter has an interconnecting function       Medicine + Technology Tübingen-Tuttlin-
and provides the basis for our compe-        gen, provide an optimal environment for
tence at the junction between micro-         the activities of the NMI.
and nanotechnology and life sciences.        The importance of tissue engineering
                                             in regenerative medicine is generally
During the past two years we extended        accepted, although this market is devel-
our growth in pharma and biotech-            oping slowly. Starting points are the de-
nology, notwithstanding the difficulties     velopment of regenerative biomaterials
of many companies in this market. This       and a better understanding of the enti-
positive development is due to our activi-   ties which influence cell differentiation.
ties in the area of functional genomics,
in which we have developed protein           Surface and interface technology,
BioChips and recombinant cell lines used     our third area of competence, plays an
by clients for target validation in the      important role in the analysis, develop-
drug discovery process. Further growth       ment and production of nano- and
is anticipated from the application of       microsystems for biosensors and neuro-
our recent development in automated          implants. One of our unique selling
electrophysiology systems and of a new       points is the scientific know-how and the
miniaturized cell biochip.                   longstanding practical experience of our
                                             staff in the area of material sciences and
The biomedical technology area repre-        surface analysis. It allows us to under-
sents a very important economical sector     stand and influence the processes which
in Baden-Württemberg. Local structures       take place at the interface between
and networks such as the German              materials and living matter.




Our areas of competence


                         Applied R&D at the interface between life- and material sciences

       Applied R&D



   Pharma and                         Biomedical Technology             Surface and
   Biotechnology                                                        Interface Technology


   Functional Genomics                Neurotechnology                   Microsystems and
                                                                        nanotechnology

                                                                                                Accredited Testing Laboratory
   Test-systems for                   Tissue Engineering                Functional surfaces     DIN EN ISO/IEC 17025.
   drug discovery                                                       and layers


   Bioanalytics/Synthesis             Development of                    Bonding, adhesion and
                                      medical products                  tribologic systems
                                                                                                DAP-PL-3420.00



                                                                                                NMI Annual Report 2002/03       5
NMI Overview
                                                                  Good news




                                                                                                              Good news in difficult times

Mio. e
7.0
                  Revenues from industrial sector                                          6.5                During the past few years there has
                                                                             6.3
6.0               Grants from public sector
                                                                                                              been a significant decrease in economic
                  Support from state Baden-Württemberg         5.3
                                                                                                              growth worldwide. This has also affected
5.0                                                                          2.5           3.0
                                                                                                              the pharma, the biotechnology and the
                                           4.3
                                                                                                              medical device industry. Despite these
4.0               3.8
                                                               2.2                                            unfavorable conditions, the NMI has
                                           1.3                                                                succeeded in the acquisition of many
                  0.9
3.0                                                                                                           new clients: revenue from industry con-
                                                                             2.1
                                                                                           2.0                tracts since 1999 has grown at a yearly
                  1.4                      1.5                 1.6
2.0                                                                                                           rate of 35%.
                                                                                                              Over the same period revenue from
1.0
                  1.5                      1.5                 1.5           1.7           1.5
                                                                                                              public funding (German ministry of
                                                                                                              education and research (BMBF),European
0.0
                 1999                  2000                   2001           2002          2003               Union (EU), state of Baden-Würt-
                                                                                                              temberg) of cooperative projects with
      Growth in revenues for the 4th consecutive year: the industrial sector grew from 0.9 to 3.0 Mio Euro.
                                                                                                              industrial partners increased at a yearly
Mio. e
                                                                                                              average of 9%. This is a positive result,
6.0
                   Surface and interface Technology
                                                                                                              in view of the fiscal restrictions during
                   Biomedical technology                                                    5.0
                                                                                                              the past years of economic stagnation.
5.0                                                                          4.6
                   Pharma and biotechnolgy
                                                                                            1.2   24%         The Ministry of Economic Affairs of
                                                                             1.0    22%
4.0
                                                               3.8                                            Baden-Württemberg was able to sup-
                                                                                                              port the NMI with 1.5 Mio Euro in the
                                                               1.1     29%                  1.2   24%
                                           2.8                               1.3    28%
                                                                                                              year 2003. This amount corresponds
3.0
                  2.3
                                                                                                              to only 24% of total revenues (6.5 Mio
                                           0.7    25%
                  0.5     22%
                                                                1.0    26%                                    Euro). Thus the negative development
2.0
                                           1.0    36%
                                                                                                              of the past four years is persisting and
                  0.8     35%
                                                                             2.3    50%
                                                                                            2.6   52%         even accelerating: while revenues from
1.0                                                            1.7     45%                                    projects have increased continuously,
                  1.0     43%              1.1    39%
                                                                                                              the percentile of funding by the regional
0.0
                 1999                      2000                2001          2002          2003               ministry is rapidly decreasing. This is a
                                                                                                              very unfortunate development, because
      Growth in revenues especially in our core competence areas pharma and biotechnology.
                                                                                                              for the NMI this support is absolutely
                                                                                                              essential in order to:1) provide R&D serv-
Mio. e
                                                                                                              ices at affordable prices to the smaller
4.0                                                                                                           enterprises, 2) perform internal R&D of
                   SME inside BW                    SME outside BW
                                                                                                              strategic importance, and 3) update the
                   large firms inside BW            large firms outside BW                                    infrastructure of the institute by invest-
                                                                                           3.0
3.0                                                                                        0.1                ing in new state-of-the-art equipment.
                                                                             2.5
                                                                             0.1
                                                               2.2                         1.2                During 2003 the NMI performed projects
2.0
                                                               0.2           0.7                              for small and medium enterprises in
                                                               0.6                                            Baden-Württemberg in the amount of
                                                                             0.2
                                           1.3                                             0.5                Euro 1.2 Mio Euro. The funding by the
                                           0.1                 0.3
                                                                                                              regional government of Baden-Würt-
1.0               0.9                      0.4
                  0.1
                                                                             1.5
                                                                                                              temberg is a prerequisite for the ability
                                           0.1
                  0.3
                  0.1
                                                               1.1                         1.2                of the NMI to support product devel-
                                           0.7
                  0.4                                                                                         opment and innovation of small and
0.0
                 1999                  2000                   2001           2002          2003               medium enterprises in this region.
      In 2003: NMI services for SME’s in the state Baden-Württemberg grew to 1.2 Mio Euro.




6     NMI Annual Report 2002/03
Steering instruments




Research institute run as a business



By the end of 2003 the NMI had 90            During the past years controlling
employees or 66 full-time equivalents.       instruments have been implemented.
There are 13 competence teams and            Operative goals are persued through
each team-leader is responsible for          management by objectives. The strategic
project acquisition, revenues, costs and     goals of the NMI are defined every two
contribution margin of his team. He          years, operative goals of each team once
reports directly to the managing director.   a year as part of the process leading to
This highly flexible horizontal organiza-    definition of the business plan of the
tion allows operations to be carried out     institute. Each team-leader has access to
in accordance with the overall strategic     all relevant data needed to run his team
goals of the institute.                      as a profit center.
                                             This organization in conjunction with the    Reinhard Rubow,
                                                                                          Chief financial officer.
Assay development                            introduction of a QM system has proven
Dr. Markus Templin, biologist                to be instrumental to our increased qual-
Bioanalytics/synthesis                       ity, efficiency and growth.
Dr. Dieter Stoll, chemist
Biochemistry
Dr. Thomas Joos, biochemist
Biophysics
Dr. Alfred Stett, physicist
Electrophysiology
PD Dr. Elke Guenther, biologist
Interface- and microtechnology
Dipl.-Phys. Werner Dreher, physicist
Molecularbiology
PD Dr. Hansjürgen Volkmer, biologist
Neurobiology
Prof. Dr. Burkhard Schloßhauer,
biochemist
Surface technology
Dr. Bernhard Schröder, physics chemist
Physical chemistry and
sensor technology
Dr. Martin Stelzle, physicist
Physical technology
Dr. Wilfried Nisch, physicist
Cellbiology
Dr. Reinout Stoop, biologist
Cellular testsystems
                                             Strategic planning
Dr. Brigitte Angres, biologist




Controlling
                                  Contribution margin
                                                                                         NMI Annual Report 2002/03   7
NMI Overview
                                                                                 Network




                                                                                 Successful R&D through networking
                                                                                 with competent partners

                                                                                 The solution of complex problems is best           The NMI is active in 27 national and
                                                                                 achieved by cooperating with compe-                international projects (cf. p. 51) involv-
                                                                                 tent partners. For this reason we define           ing over 100 partners and a volume of
                                                                                 and coordinate collaborative national              48 Mio Euro. The NMI is project leader
                                                                                 and international projects in which we             in 12 such projects with a volume of 28
                                                                                 bring together partners from academic              Mio Euro. A large part of these revenues
                                                                                 and non-academic institutions, research            is to the benefit of small and medium
                                                                                 foundations, industry, clinics, etc.               enterprises and of our partners at the
                                                                                 Through this interdisciplinary network             University of Tübingen.
                                                                                 we achieve complementation of know-
                                                                                 how and allow our partners to benefit
                                                                                 from the entire value chain, i.e. from
                                                                                 basic research to product to market.




                                                                     Universität Stuttgart
                                                                                             Universität Oulu,
                                                                                             Finnland


                                                                                              FH IPM Freiburg                Heinrich-Heine-Universität
                                                                                                                             Düsseldorf
                           Universität Tampere,
                           Finnland
                                                                                      Philipps-Universität                    Ulm       INI Hannover

Universitätsklinikum                                                                  Marburg
Würzburg

                                                                                        Universität Zürich


                                                                   LMU München
    Villingen-Schwenningen                                                                                                Leopold-Franzens Universität
                                                                                                                          Innsbruck


Universität zu Köln                                                                      Biozentrum Würzburg                IMTEK Freiburg

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                                                                                             Ruprecht-Karls-Universität
                                                                  FH Albstadt
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                                                                                             Heidelberg
                                                                  Sigmaringen
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      Universitätsklinikum Ulm                                                                                                            Denkendorf
                                                                                                       Unversität Würzburg


                                                            FH Reutlingen                    Universität Kaiserslautern

                                           Friedrich Schiller Universität
                                           Jena                                                                                IMS Stuttgart
                                                                                                  Stiftung Orthopädische
                                                                      Universitätsklinikum        Universitätsklinik Heidelberg
                                                                      Regensburg




8   NMI Annual Report 2002/03
Start-up Companies




NMI as incubator for new                                              The NMI TT GmbH,
enterprises                                                           founded in the year 2002

New enterprises are founded at sites                                  The demand for finished products and
which provide the best environment                                    services based on established technolo-
to the entrepreneur. The NMI has pro-                                 gies can only partially be met by the
vided a technological basis to numerous                               NMI, which is more focused on applied
new start-up companies and cooperates                                 R&D. The NMI Technology-Transfer (NMI
with them in R&D projects supported                                   TT) GmbH offers products and services
through public funding. Many of these                                 based on technologies developed at the
new enterprises utilize our know-how,                                 NMI. High quality at competitive prices
skilled personnel, infrastructure and                                 are distinguishing features of the NMI TT
equipment to perform their R&D and                                    GmbH.
product development, thus reducing
their need for capital investment, per-                               The NMI TT GmbH is a 100% subsidiary
sonnel and minimizing financial risks.                                of the NMI, founded to commercialize
This incubator concept has proven to be                               NMI know-how:
very successful.                                                      • IP protection and exploitation
Another important factor for new start-                               • Peptide synthesis
up companies is the strong commitment                                 • Bioanalytics
and engagement of the cities Tübingen                                 • Analysis of materials
and Reutlingen (founders of the TTR                                   • Coatings
Technology Park Tübingen-Reutlingen),                                 • System analysis, technology
of the local Chamber of Commerce and                                    assessment and consulting
of the University of Tübingen.
As an example for this favourable envi-
ronement one can mention the federal
funding (BMBF) of projects in the area of
regenerative biology through the initia-
tive “Bioprofile”. Through this program
the region Stuttgart-Tübingen-Esslin-
gen-Reutlingen has obtained funding
in the order of 15 Mio Euro for projects
in which small and medium enter-
prises together with academic or other
research institutes (e.g. the NMI) pursue
the development of new products
related to regenerative biology.

                                                                                    NMI TT GmbH
             Knowledgeable, courageous and
             engaged researchers took up
             technolgies and created enterprises.
             These new start-up companies
             cooperate with the NMI and the
             University of Tübingen.




                                                    accelab
                                                          Retina Implant AG




                                                                                     NMI Annual Report 2002/03   9
NMI Overview
                                             Patents




                                             Patents - intellectual property for
                                             the protection of innovation

                                             A patent-portfolio is the most valuable      of intellectual property to our clients in
                                             asset of any technology company and          exchange for license fees has been an
                                             research institution.                        important incentive for our clients to
                                             To develop and secure intellectual           cooperate with the NMI. Since 2003 the
                                             property, the NMI offers its employees       NMI transfers all its intellectual property
                                             incentives which exceed compensation         to the NMI TT GmbH, which pursues
                                             mandated by law. During the past years       patent commercialization. In many cases
                                             the NMI has made significant progress        NMI patents are transferred to spin-off
                                             in establishing a patent portfolio, with     companies in return for a small participa-
                                             more then 40 patent families filed or        tion in the company.
                                             assigned. In many cases employees
                                             of the NMI are listed as inventors in
                                             patents of our clients. The NMI spends
                                             approximately 3% of its income for
                                             patent filing and protection. The transfer




Numerous patents secure the IP of the NMI.




10   NMI Annual Report 2002/03
Public relations




Marketing and public relations



The NMI is visible through its presence          •   Over 80 oral and poster presentations            Contact:
at exhibitions, presentations at scientific          at scientific conventions in 2003                Dr. Nadja Gugeler
congresses, publications in scientific           •   Teaching at Universities                         phone: +49 7121 51530-842
journals and articles in the daily press.        •   Advising graduate and undergraduate              gugeler@nmi.de
The most important and effective public-             students performing their thesis at
ity, however, is the reference from our              the NMI
satisfied clients: good quality recom-           •   Introduction of high-school students to
mends itself. In today’ s competition                scientific work through their participa-
among contract research organizations,               tion in research projects at the NMI
the international reputation of our scien-
tific staff is not sufficient to gain market
share. Our new ideas, activities, results
and success cannot by themselves attract
the interest of our potential clients. Vis-
ibility has therefore become an essential                                                             With support of the Ministry for Science and Art of
component of our marketing strategy.                                                                  the state Baden-Württemberg the cluster “BioChip-
                                                                                                      technologies“ was formed. As part of this project
                                                                                                      the databank www.biochipnet.de has been estab-
•   Internet: www.nmi.de, homepage;                                                                   lished. In 2003 the site was visited 98.000 times.
    www.biochipnet.de, databank                                                                       Dr. Jutta Bachmann, Bachmann Consulting, Norway,
                                                                                                      in cooperation with the NMI is responsible for the
•   TV, radio: film, interviews and reports
                                                                                                      updating of the databank.
•   Print media: product sheets, yearly
    report, press releases
•   Exhibitor at fairs : Analytica, Bioanalyt-
    ica, BioDigital, Biotechnica, Hannover
    Messe, Medica, MEDTEC
•   Organization and execution of scien-
    tific meetings, seminars
•   Publications in scientific journals




                                                                      N. Gugeler is responsible for
                                                                      public relations.




                        “Simply
                              the best!”
                                                                                                                        NMI Annual Report 2002/03      11
NMI Overview
                                       Meeting clients’ expectations




                                       Customer satisfaction, key to success



                                       What does our client need?                   “A poll among 40 clients gave a very
                                       or                                           positive result, with 70 percent return
                                       Which client will need our technology?       of questionnaires. Eighty percent were
                                                                                    extremely satisfied and twenty percent
                                       These are two central questions for the      were very satisfied with the quality of
                                       provider of R&D services. For the NMI a      our services, in particular with respect to
                                       difficult act of balance between meeting     technical and scientific competence of
                                       the current market needs and anticipat-      the project leader, technology offerings,
                                       ing future technological and market          execution of contract in time and costs,
                                       developments.                                content and quality of the final report.
                                       For our employees is it also a question      We thank our clients for their confidence
                                       of self-image, having to fulfill the dual    and, with their critical input and support,
                                       role of scientist and service provider.      we will continue to improve the quality
E. Müller is pleased with the poll     They have succeeded in bringing              of our work”.
confirming a high degree of customer   together these two aspects. With the
satisfaction.                                                                               Enzio Müller, Ph.D., Managing Director
                                       introduction of the QM-system in the
                                       year 2000 our scientists have become
                                       more aware of their role as service
                                       providers focused on meeting clients’
                                       expectations. Through our public relation
                                       and marketing activities, we will increase
                                       our visibility and extend the radius of
                                       action into international markets, while
                                       continuing to support the regional small
                                       and medium enterprises.




                       Quality management
Customer satisfaction
                                                                        Public relations


12   NMI Annual Report 2002/03
Testing according to DIN EN ISO/IEC 17025




Testing of medical products in accredited
areas according to DIN EN ISO/IEC 17025

Current legislation mandates that the
testing of medical products for registra-
tion must be performed in accredited
laboratories. This can also be the case
for industrial products. A prerequisite
for accreditation is the establishment
of a quality management system. The
latter has been introduced at the NMI.
All services provided to our clients are
performed according to the QM system.
Testing can be performed in accredited
areas according to the European norm
EN ISO/IEC 17025.




                                            Document of the accreditation by the DAP.




                                                                           DAP-PL-3420.00




Quality management screenshot.




                                                                                        NMI Annual Report 2002/03   13
Pharma and biotechnology



                                     Pharma and biotechnology



                                     In cooperation with pharma and bio-
                                     technology companies the NMI develops
                                     new procedures and system solutions for
                                     drug discovery.
                                     The sequencing of the human genome
                                     represents a milestone in modern biol-
                                     ogy. For the pharmaceutical industry it
                                     opens many new opportunities for the
                                     identification of specific disease-associ-
                                     ated genes and cellular targets. This will
                                     facilitate the drug discovery process and
                                     lead to the treatment of many diseases.
                                     The major effort in the post-genomic era
                                     will reside in the identification, func-
                                     tional characterization and comparative
                                     analysis of all cellular proteins (so-called
                                     proteom) in healthy and diseased tissue.

                                     We work in the areas:
                                     • Functional genomics
                                     • Test-systems for drug discovery
                                     • Bioanalytics/peptide synthesis




                           Among our customers are:
                                     Abbott Laboratories, Illinois/USA
                                     Accelab GmbH, Kusterdingen
                                     Advalytix AG, Brunnthal
                                     Alopex GmbH, Kulmbach
                                     Altana Pharma AG, Konstanz
                                     Bachmann Consulting, Nesoddtangen/
                                     Norwegen
                                     Bayer AG, Leverkusen
                                     BMI Biomedical Informatics,
                                     Heidelberg
                                     Cytocentrics CCS GmbH, Reutlingen
                                     Grünenthal GmbH, Aachen
                                     Ingenium Pharmaceuticals AG,
                                     Martinsried/Planegg
                                     Invitrogen Corporation, Carlsbad, CA/USA
                                     Merck KgaA, Darmstadt
                                     Merz Pharma GmbH & Co. KgaA, Frankfurt
                                     MorphoSys AG, Martinsried/Planegg
                                     Multi Channel Systems MCS GmbH,
                                     Reutlingen
                                     N-Zyme BioTec GmbH, Darmstadt
                                     Schering AG, Berlin
                                     Servier, Neuilli-Sur-Seine Cedex/France
                                     TaiMont Biotech Inc., Tainan/Taiwan
                                     UniLever, Vlaardingen/Holland
                                     Zeptosens AG, Witterswil/Schweiz
Functional genomics                            Test-systems for drug discovery              Bioanalytics/Synthesis



From gene to protein to cell function          In vitro technology platforms for the        Custom-specific solutions with claim
                                               post-genomic era.                            to quality

Gene transfer                                  Electrophysiology with tissues               Bioanalytics
With viral vectors we introduce gene           and cells                                    Highly sensitive and specific analytical
sequences of putative disease-relevance        With microelectrode arrays (MEA tech-        methods for research and quality control
into cells. This allows induction or inhibi-   nique) one can measure and analyze the       require expensive instrumentation and
tion of expression of proteins and assess-     propagation of signals upon electro-         expert personnel. The NMI provides cus-
ment of their role in a given disease          stimulation as well as the spontaneous       tomers with specific problem solutions
(p. 16-17).                                    activity of electrogenic cells (p. 21-23).   (p. 26).

Protein BioChips                               Characterization of ion channels             Peptide synthesis
A large number of proteins from a small        Messenger RNA or cDNA coding for ion         Peptides play an important role in
tissue sample can be identified simul-         channels is injected into frog Xenopus       research and clinical medicine. We offer
taneously and with high sensitivity in a       laevis oocytes. The ion channel proteins     rapid, custom-specific synthesis of pep-
miniaturized assay at low cost (p.18-19).      are expressed and after one day drug         tides at competitive prices (p. 27).
                                               effects can be measured by a robot
Cell BioChips                                  (p. 23).
The proteins of the extracellular matrix
influence cell function. We have devel-        With a new microsystem technology
                                       TM
oped a multiple substrate array (MSA )         approach developed by the NMI with
for the screening of cell-matrix interac-      industrial partners, automated patch-
tions (p. 20).                                 clamp recording becomes a reality.

                                               Blood-Brain-Barrier
                                               With partners from pharmaceutical
                                               industry the NMI has developed an
                                               in vitro model of the blood-brain-barrier
                                               (p. 25).




Our services for drug discovery support

                                                          Protein-BioChips
              Gene transfer                                  CellBioChips                            Blood-Brain-Barrier
                                                                                Electrophysiology
                                                          Peptide synthesis                              Bioanalytics


       Identification         Validation of        Assay                  Screening of        Pharmacology          Toxicology
       of targets             targets              development            substances



Pharma value chain: preclinical phase




                                                                                                           NMI Annual Report 2002/03   15
Pharma and biotechnology
                                                       Functional genomics




                                                      Gene transfer



Contact:                                              Transgenic cells - a starting point for       “We are experts in developing highly
PD Dr. Hansjürgen Volkmer                             drug discovery                                efficient procedures for the establish-
phone: +49 71 21 5 15 30-44                                                                         ment of cell-lines for target validation.
volkmer@nmi.de                                        Transgenic cells are used in the drug         As a service to pharmaceutical industry,
                                                      discovery process for target identification   we optimize and apply molecular biol-
Dr. Frank Weise                                       and validation.                               ogy techniques to study the function
phone: +49 71 21 5 15 30-56                           Putative disease-related genes are identi-    of proteins of specific interest to our
weise@nmi.de                                          fied by comparative genome analysis of        customers. We have developed adeno-
                                                      healthy and diseased humans.                  viral vectors for the endogenous siRNA
                                                      These genes can be transferred into cells     formation. This allows to inhibit the
                                                      via viral vectors. This leads to transient    expression of specific proteins in primary
                                                      or stable over-expression of the gene         cells. On the other hand, we can induce
                                                      product, i.e. the protein. Conversely,        overexpression of proteins through viral
                                                      by inducing siRNA one can inhibit the         vector mediated gene transfer. With
                                                      expression of specific proteins. With this    these tools we can study the role specific
                                                      approach it is possible to analyze the        proteins play in the modulation of cel-
                                                      function of proteins (potential drug tar-     lular processes such as differentiation,
                                                      gets) in their cellular context and draw      apoptosis and cell vitality. Such data are
                                                      conclusion with respect to the role of        important for drug discovery in order
                                                      specific proteins in a given disease.         to identify new targets for therapeutic
                                                                                                    intervention.”
                                                                                                                       PD Dr. Hansjürgen Volkmer,
                                                                                                                        Head of Molecular-biology




                                                                                                    The high number of human genes,
                                                                                                    their mutations, the variants of their
                                                                                                    expression products, i.e. proteins, and
                                                                                                    the differences from tissue to tissue by
                                                                                                    which clusters of proteins regulate cell
                                                                                                    function, reveal the complexity of the
                                                                                                    task faced by researchers in the pharma-
                                                                                                    ceutical industry in their effort to identify
                                                                                                    specific targets linked to a disease.




                    From the left: HJ. Volkmer, F. Weise and S. Sturany.




16   NMI Annual Report 2002/03
In order to analyze in parallel the func-
tion of many genes, the NMI has devel-
                            TM
oped a chip (AdenoChip ) on which
adenoviruses carrying different genes are
spotted as 100-dot arrays. When cells
                                 TM
are applied to the AdenoChip gene
transfer takes place and the effect of                                         A                                    B
gene expression on cell function at each
position on the array can be observed.
This allows the simultaneous analysis of
the function of genes and their expres-
sion products, i.e. proteins, thereby
facilitating target identification for drug
discovery.

Facilities/methods
• S1/S2 laboratories
• Confocal laser scanning microscope
  (Zeiss)                                                                      C
• Fluorescence microscopes,
  Image analysis systems
• Luminometer (Dynex Technologies)
• FLUOstar OPTIMA Plate-reader
  (BMG Labtechnologies)
• Quantitative RT-PCR (TaqMan)


•   Expression of recombinant proteins in
    pro- and eucaryotic systems.
•   Viral gene transfer (adenovirus and
    retrovirus)
•   Neuronal growth and regeneration
    in vitro
•   Inhibition of gene expression
    (siRNA/shRNA)
•   Protein-BioChips
•   Reporter assays (luciferase, green
    fluorescent protein)




                       Silencing of the neurofascin gene achieved by shRNA-encoding adenoviruses
                       Chick tectal neurons were cultivated on neurofascin as a substrate. A: Neurons infected by a control adenovi-
                       rus. B: Neurons infected by an adenovirus expressing shRNA directed against neurofascin (bar: 100 µm).
                       C: Determination of the percentage of neurons with neurites. The outgrowth of neurites observed after
                       infection by adenoviruses expressing shRNA directed against neurofascin is significantly reduced compared
                       to the controls, in which cells were either not infected at all, or in which cells were infected by a control
                       adenovirus.




                                                                                                                               NMI Annual Report 2002/03   17
Pharma and biotechnology
                                                         Functional genomics




                                                         Protein-BioChips



Contact:                                                 Chips to Hits                                  tions, conferences, etc. can be found
Dr. Thomas Joos                                                                                         in our databank - www.biochipnet.de -
phone: +49 71 21 5 15 30-844                             ... is the title of an international confer-   which is being constantly updated. In
joos@nmi.de                                              ence on BioChip applications for drug          2003 there were up to 98.000 visits per
                                                         discovery and diagnostics.                     month (p. 11).
Dr. Markus Templin
phone: +49 71 21 5 15 30-828                             BioChips allow the simultaneous analysis
templin@nmi.de                                           of thousands of molecules extracted
                                                         from minute samples of tissue or body
                                                         fluids. This is done by applying on
                                                         specially coated slides under clean-room
                                                         conditions specific capture molecules as
                                                         arrays of minispots using instruments          NMI, a pioneer in Protein-BioChips
                                                         called arrayers. Analytes in the sample
                                                         bind to their respective capture mol-          In the blood of patients with autoim-
                                                         ecules and quantitative analysis of bind-      mune diseases one can detect autoan-
                                                         ing parameters is performed by optical         tibodies. With protein-microarrays
                                                         procedures.                                    (BioChips) one can quantify these
                                                                                                        autoantibodies with the same sensitiv-
                                                         The field of BioChips has rapidly              ity and specificity as with conventional
                                                         expanded in the past five years, leading       ELISA-assay, but with much less re-
                                                         to a great variety of instruments, tech-       agents and sample specimen. BioChips
                                                         nologies and applications. An overview         of proteins find their application also
                                                         of companies, technologies, publica-           in the measurement of signal molecules
                                                                                                        (cytokines), proteins related to cell dif-
                                                                                                        ferentiation and in antibody screening
                                                                                                        (Templin, M. et al. 2003; Stoll, D. et al.
                                                                                                        2004).

                                                                                                        “We have focussed early on the devel-
                                                                                                        opment of protein-BioChip microar-
                                                                                                        rays for the measurement of antigens,
                                                                                                        antibodies, enzymes and other protein
                                                                                                        molecules. The interest of industry for
                                                                                                        miniaturized and parallelized assay
                                                                                                        systems is very high. However, there are
                                                                                                        still significant obstacles in the area of
                                                                                                        automation which need to be overcome
                                                                                                        before a breakthrough of the protein-
                                                                                                        BioChip technology can occur”.

                                                                                                                Dr. Thomas Joos, Head of Biochemistry




From left to right: B. Angres, M. Templin, T. Joos and D. Stoll: biologists,
biochemists and chemists discuss applications of the Cell-BioChip (MSATM).




18   NMI Annual Report 2002/03
PepART: Peptide based protein                 Assay development
analysis with Array Technology

Antibody microarrays allow simultan-          The NMI has invested considerable                        Equipment
eous measurement of many proteins or          amounts of money to equip the labora-                    Cleanroom for BioChip production with
peptides in complex samples. Such             tories with the most advanced instru-                    • Perkin-Elmer Arrayer BCA1
a system would be very useful for pro-        mentation needed for the development                     • Genetic-Microsystems 417
teome analysis. However, to generate a        of miniaturized and parallelized assays.
large number of specific antibodies by                                                                 BioChip reader
conventional approach, one would need         ”Our most modern instrumentation and                     • Zeptosens reader
to isolate a correspondingly high number      our technical and scientific know-how                    • Genetic-Microsystems 418
of antigens. The PepART approach              enable us to offer solutions tailored
circumvents this difficult task by using      to the specific needs of each client.                    Bead-based System
synthetic peptides as antigens, whereby       While planar microarry chips are useful                  • Luminex100
the peptides are selected based on            for many applications, we see a great                    • Luminex HTS
published protein sequence data, such         potential in the bead-based technologies
that the peptide sequence (so called          which allow simultaneous measurement                     Automation
signature peptide) is present only in         of up to 100 analytes in a single probe.                 • Quiagen BioRobot 3000
the protein of interest and not in other      When extreme sensitivity is required, we                 • Tecan Genesis
proteins composing the proteome. With         use the “planar waveguide” technology
such antibody arrays specific proteins in     of Zeptosens.”
complex samples can be detected upon
                                                                           Dr. Markus Templin,
proteolytic fragmentation of the sample.
                                                                   Head of Assay Development
This research project (FKZ0312879A) is
funded by the BMBF (federal ministry
for education and research). Project
partners are scientists at the Universities
of Tübingen, Ulm and Zürich, and at
the companies MorphoSys AG, Accelab
GmbH, Merck KGaA, Ingenium AG and
BMI GmbH.



Microfluidic processor for diagnostics

A lab-on-the-chip-system for sample
preparation and processing is being
developed at the NMI. It will integrate
the transport, mixing and dispensing
of minute liquid aliquots under defined
reaction condition. The biotech compa-
nies Advalytix AG, Alopex GmbH and the
NMI are partners in this project funded
by the BMBF (FKZ 16SV1769).
                                              Peptide microarrays for the determination of epitope specificity and crossreactivity of antibodies.
                                              Recombinant antibody fragments enriched on a linear peptide of 25 aminoacids were analyzed for their
                                              binding properties using peptide microarrays. Arrays consisting of different variants of the initial antigen
                                              were created and incubated with purified antibody fragments. The picture shows binding on systematically
                                              mutated peptides (red frame), full length peptide (white), C-terminally shortened (green) and N-terminal
                                              shortened (magenta) variations of the antigen; included controls are printing buffer (grey), anti Fab IgG
                                              (blue), BSA (orange) and a set of negative control peptides (yellow). The amount of antibody captured on a
                                              spot is quantified and used to define the epitope specificity and crossreactivity of the binder.




                                                                                                                          NMI Annual Report 2002/03          19
Pharma and biotechnology
                                 Functional genomics




                                 A new Cell-BioChip


                                                                                                                 TM
Contact:                         Proliferation, adhesion, migration, dif-   In September 2003 the MSA -system
Dr. Brigitte Angres              ferentiation and apoptosis of cells are    was presented at an international con-
phone: +49 71 21 5 15 30-838     influenced by the extracellular matrix.    vention in the USA. Dr. B. Angres, Head
angres@nmi.de                    Thus, cell-matrix interactions regulate    of Cellular Assay Systems at the NMI,
                                 organogenesis, influence tumor and         was awarded the “Best poster award”
                                                                                                          TM
                                 metastasis formation, and promote heal-    for the originality of the MSA concept.
                                 ing and regeneration of injured tissues.
                                                                            “The Multi-Substrate-Array can be
                                 The functional variety of extracellular    used for rapid screening of the effects
                                 matrices results from specific combi-      of matrix molecules on cell function in
                                 nations of diverse classes of matrix       regenerative medicine, cancer and bio-
                                 molecules.                                 material research. In 2004 we will begin
                                 We developed the Multiple Substrate        beta-testing with selected partners.”
                                             TM
                                 Array (MSA ) for the parallelized and
                                                                                                          Dr. Brigitte Angres,
                                 low cost analysis of effects of many                                Head of Cellular Systems
                                 matrix components and mixtures thereof
                                 on cells.




                                                                            The MSA™ consists of arrays with 64 spots of dif-
                                                                            ferent extracellular matrix molecules with a diameter
                                                                            of 300 µm. Eight of those arrays are spotted on one
                                                                            slide and can be separated from each other by a
                                                                            removable chamber system. Cells can be seeded in
                                                                            the chambers and their responses to extracellular
                                                                            cues can be analyzed after hours or days in a mini-
                                                                            aturized and parallelized way.




20   NMI Annual Report 2002/03
Test systems for drug discovery




Electrophysiology: innovations
through microtechnology

Defects of ion channel function and                      The NMI develops new systems for the                  Contact:
faulty signal transduction are the culprit               automated measurement of electro-                     Prof. Dr. Elke Guenther
for many diseases, such as diabetes, car-                physiological parameters, with the aim                phone: +49 7121 51530-54
diovascular and nervous system diseases.                 of providing “screening tools”.                       guenther@nmi.de
Electrophysiological methods are used in
basic and clinical research                              “Our goal is to introduce automation
to understand the molecular basis of                     in electrophysiology. Our scientists in
many diseases and to develop new                         the area of microtechnology, biophys-
drugs. Electrophysiological studies                      ics, neurobiology and electrophysiology
require highly skilled personnel and                     collaborate with the companies Multi
are time consuming. Therefore in                         Channel Systems MCS GmbH and with
pharmaceutical industry they are more                    Cytocentrics CCS GmbH.
frequently used in the more advanced                     For our clients in the pharmaceutical
phases of drug discovery.                                industry we perform electrophysiologi-
                                                         cal measurements in vitro both for drug
                                                         discovery and for safety pharmacology
                                                         studies.”
Microelectrode array (MEA) developed by the NMI
for electrical stimulation and recording of electrical                           Prof. Dr. Elke Guenther,
activity of living tissues. The amplifier contact pads                         Head of Electrophysiology
to the 60 electrodes in the centre of the incubation
chamber are aligned along the chip sides.

  1 cm




                                                                                                               E. Guenther and B. Schloßhauer.




200 µm                                                       10 µm                                             200 nm

Brain slice (hippocampus) on a microelectrode array          A single titanium nitride electrode. The highly   The nano-structure of the pin-like titanium nitride
(MEA). The electrical activity can be recorded from          sensitive and electrodes with long-term stabi-    provides a large surface, a prerequisite for the elec-
60 electrodes simultaneously thus allowing the               lity are manufactured at the NMI (p. 40).         trodes´ outstanding charactristics.
analysis of the spatiotemporal signal propagation
within the brain slice.



                                                                                                                                   NMI Annual Report 2002/03        21
Pharma and biotechnology
                                 Test systems for drug discovery




                                 Electrophysiology with cells and tissues



Contact:                         Measurements with microelectrode                   MEA Cardiosensor
Prof. Dr. Elke Guenther          arrays (MEA)                                       ECG from cell cultures
phone: +49 71 21 5 15 30-54
guenther@nmi.de                  Microelectrode arrays on glass support             A potential adverse effect of many
                                 are produced in the clean room by use of           drugs is the induction of life-threatening
Dr. Ulrich Pehl                  photolitographic techniques (p.40). They           cardiac arrhythmia in patients already
phone: +49 71 21 5 15 30-896     are used for the measurement of field              suffering from cardiovascular diseases.
pehl@nmi.de                      potentials and for the electric stimulation        This is recognized in the ECG through
                                 and recording of activity of electrogenic          prolongation of the QT-interval. The
                                 tissues or groups of cells. As the leader          guidelines of the International Confer-
                                 in the field of MEA technology, we pro-            ence on Harmonization (ICH) prescribe
                                 duce arrays of different geometry and              that drugs for use in humans are tested
                                 electrode densities for our partner, the           in safety pharmacology studies for their
                                 company Multi Channel Systems MCS                  potential to prolong the QT-interval.
                                 GmbH (www.multichannelsystems.com).                We use the MEA Cardiosensor as a rapid
                                 This product is the result of a project            test-system which fulfills the ICH require-
                                 carried out in cooperation with research-          ments. We determine for our clients
                                 ers at the University of Freiburg and              the effect of drugs on the duration of
                                 Köln and the companies Multi Channel               extracellular field potentials (QT-interval)
                                 Systems MCS GmbH and Medigene AG                   in cardiac tissue.
                                 with funding from the BMBF
                                 (FKZ 0310964A).




                                                        Upper picture:
                                                        Whole heart preparation of embryonic chicken
                                                        on a MEA.

                                                        Lower panel:
                                                        Drug effects on field action potential (fAP)
                                                        prolongation.
                                                        Left: Dose-response effect of the antiarrhythmic
                                                        drug quinidine on the fAP duration.
                                                        Right: Dose-response curves of the effects of the
                                                        antiarrhythmic drugs quinidine and sotalol and
                                                        the HERG channel blocker E4031 on the fAP
                                                        duration of ventricular myocytes.



22   NMI Annual Report 2002/03
                                                                                                 Characterization of ion channels



MEA Retinasensor                          MEA Neurosensor                                        Roboocyte
Retina function in vitro                  Coupling nerves to chips                               Rapid screening through automation

Disturbances of the visual system are     For the therapy of spinal chord injury,                Injection of mRNA or cDNA in frog
a potential side-effect of many drugs.    stroke and other neurodegenerative dis-                oocytes can be used to induce the
For this reason new therapeutic agents    eases one would like to discover drugs                 synthesis and membrane insertion of
must be tested to exclude such adverse    which induce functional neurite growth                 ion channels. With appropriate elctrodes
effects.                                  (as opposed to random non-functional                   one can record the electrophysiological
Retina explants can be placed on MEAs     growth). For this reason the neuropro-                 activity and the effect of compounds
to record micro-electroretinograms        tective and regenerative potential of                  on these channels. For rapid and cost-
(microERG). This system allows to study   compounds must be tested in a model                    effective screening of the effect of
the effect (dose-response analysis) and   which resembles the in vivo situation.                 drugs on specific ion channels, we use
the mechanism of action of compounds                                                             the automated system Roboocyte of
on the neuronal network of the retina.    Our MEA neurosensor consists of an                     the company Multi Channel Systems
                                          incubation chamber placed on a MEA                     (www.multichannelsystems.com). This
                                          (p. 21). This allows to culture brain tissue           robot injects the nucleic acid, adds the
                                          for up to six weeks with intermittent                  compound of interest and performs the
                                          electrophysiological recordings. Selected              electrophysiological recording.
                                          neuronal pathways in brain slices are
                                          transected and drug mediated induc-
                                          tion of functional regeneration can be
                                          monitored electrophysiologically.




                                          Chicken or rats retinae are placed on MEA elec-
                                          trodes with the ganglion cell layer contacting the
                                          MEA. Light-evoked ERGs can be recorded from up
                                          to 60 microelectrodes.


                                          Drug actions can be studied by the analysis of         Oocytes are dissected from anesthetized african
     Light stimulus
                                          changes in the characteristics of the light-evoked     clawed frogs (Xenopus laevis). The cells are then
                                          ERGs. The figure shows the reversible elimination of   injected with mRNAs or cDNAs coding for ion chan-
                                          the b-wave by application of AP4, a blocker of the     nels. After an incubation period the induced currents
            b-wave
                                          on-signal pathway in the retina.                       are recorded and analyzed by a robot.




 AP4 100µM




  wash


                           50 µV√


                           500 ms




                                                                                                                    NMI Annual Report 2002/03      23
Pharma and biotechnology
                                                     Test systems for drug discovery




                                                     Automated Patch-Clamping



Contact:                                             Chip instead of micro-pipette                         We have produced glass-chips with
Dr. Alfred Stett                                     A breakthrough in patch-clamping                      micrometer openings which allow
phone: +49 71 21 5 15 30-70                                                                                gigaseal cell contacts similar to those
stett@nmi.de                                         The patch-clamp method allows the                     obtained with glass pipettes. Subse-
                                                     electrophysiological analysis of ion                  quent work allowed the development
                                                     channels. For this discovery Erwin Neher              of a device in which in a two step
                                                     and Bert Sackmann were awarded the                    procedure (so-called cytocentering) cells
                                                     Nobel price. The patch-clamp proce-                   are automatically positioned onto the
                                                     dure involves the use of a glass pipette              chip openings and an electric contact is
                                                     with an opening in the order of a few                 established. The company Cytocentrics
                                                     micrometers. Under microscope exami-                  AG (www.cytocentrics.com), a spin-off
                                                     nation the pipette is applied to the cell             of the NMI, has further developed the
                                                     membrane and, under gentle suction, a                 system and the first instruments for
                                                     tight seal (gigaseal) is formed between               automated patch-clamping have been
                                                     pipette and cell membrane. Further                    produced and will undergo testing at
                                                     suction removes the small patch of cell               several beta-sites.
                                                     membrane at the inside of the pipette
                                                     opening, while the cell membrane                      “The NMI is a technological partner of
                                                     outside the opening adheres to the                    Cytocentrics AG for the development
                                                     outer pipette surface. With this set-up               of an automated patch-clamp instru-
                                                     electrophsiological measurements can                  ment. Furthermore, molecular biologists
                                                     be performed and the effect of drugs on               and electrophysiologists at the NMI will
                                                     ion channels is analyzed. The procedure               assist Cytocentrics AG and pharmaceu-
                                                     requires skilled personnel and is very                tical companies in the development
                                                     time-consuming and costly. To overcome                of validated application protocols for
                                                     these limitations we have collaborated                compound screening, using genetically
                                                     with Bayer AG and invented a procedure                engineered cells.”
                                                     for the development of an automated
                                                                                                                    Dr. Alfred Stett, Head of Biophysics and
                                                     process (Stett, A., Burkhardt, C. et al.                                 co-founder of Cytocentrics AG
                                                     2003; Stett, A., Bucher, V. et al. 2003;
                                                     Knott, T. et al. 2002).




                                                     Cytocentering Technique and CytoPatchTM-Chip




                         Two independent channels are used: The cytocentering chan-       EM image of the chip (bar 1 µm)   View of the internal structure
                         nel is for suction and positioning of cells and the CytoPatch™                                     of the CytoPatch™-Chip.
                         channel is used for patch clamping and recording.                                                  Cytocentering channel 1 and
                                                                                                                            CytoPatchTM channel 2




24   NMI Annual Report 2002/03
A model for Blood-Brain Barrier studies



The blood-brain-barrier (BBB) consists       “The pigmented epithelium of the eye                   Contact:
of a network of capillaries providing        has similar brain barrier function as the              Prof. Dr. Burkhard Schloßhauer
a blood-brain exchange surface of            endothelium of the capillaries in the                  phone: +49 7121 51530-20
100 m2. It provides a constant milieu        BBB. It protects the neuronal network                  schlosshauer@nmi.de
for the brain cells by mediating selective   of the retina by excluding certain toxic
entry of substrate molecules and oxygen      blood-borne substances, while allowing
from the blood compartment and exit          entry of molecules, such as nutrients
of metabolic products and C02 from           and vitamins. Many proteins which are
the brain compartment. Furthermore, it       intrinsic to the function of the BBB are
prevents the entry of many neurotoxic        also present in the blood-eye-barrier. The
compounds into the brain. The NMI has        pigmented epithelium is prepared from
established a BBB model in cooperation       the eye of the pig as a thin sheet. The
with five German pharmaceutical com-         model has been validated as a surrogate
panies and with the support from the         for the Blood-brain-barrier.”
BMBF (FKZ 0311538) for the testing
                                                            Prof. Dr. Burkhard Schloßhauer,
of drug permeability through the BBB.                                 Head of Neurobiology
At present the only in vitro system
for BBB studies consists of confluent
endothelial and glia cells. The system
is complex, requires considerable cell-
culture experience, it is time-consuming
and costly. Our model consists of an
                                                                      Model of the microvessel system of the human brain. The capillary network has a
intact eye-brain-barrier freshly prepared                             surface of approximately 100 m².
from pig eyes which can be immediately
used for the testing of compounds.




                                                     The pigmented epithelium of the eye serves
                                                     the same function as the blood-brain barrier
                                                     (BBB) and can be used as BBB-model.


                                                                                                                    NMI Annual Report 2002/03      25
Pharma and biotechnology
                                                  Bioanalytics and synthesis




                                                  Bioanalytics



Contact:                                          Highly sensitive methods for                     Our services include:
Dr. Dieter Stoll                                  the analysis of biomolecules                     • Identification and quantification of
phone: +49 71 21 5 15 30-811                                                                         natural products (proteins, peptides,
stoll@nmi.de                                      Highly sensitive and selective analytical          oligosaccharides, phospholipids...)
                                                  procedures require sophisticated instru-         • Characterization of post-translational
Dr. Andreas Jaworski                              mentation and highly skilled personnel.            modification of proteins.
phone: +49 71 21 5 15 30-829                      We develop custom-specific solutions for         • Quantitative analysis of biomolecules,
jaworski@nmi.de                                   our clients. The results of our studies are        drugs and drug metabolites in tissue
                                                  presented in a format which meets the              extracts, body fluids, breath conden-
                                                  requirements of the regulatory bodies,             sates, etc.
                                                  e.g. FDA. The analytical methods and             • Analysis of identity and purity of drugs
                                                  procedures which we develop for spe-               produced by biotechnological proce-
                                                  cific assays are performed according to            dures (e.g. recombinant proteins).
                                                  DIN EN ISO/IEC 17025.                            • Assessment of cleanliness and effec-
                                                  Routine measurements are offered by                tiveness of medical devices ( e.g.
                                                  the NMI Technology Transfer GmbH                   pumps, drug delivery systems, drug
                                                  (www.nmi-tt.de).                                   coated stents) by quantitative analysis
                                                                                                     of contaminants and drug in fluids
                                                  “Our strength is the development and               delivered by the device.
                                                  validation of specific analytical proce-
                                                  dures to meet the specific needs of each         Equipment:
                                                  client. Quality, rapid response and data         • ESI/Apcl-qTOF mass spectrometer
                                                  delivery at competitive prices are our           • ESI/Apcl-Tripel-Quadrupol mass
                                                  distinctive feature.”                              spectrometer
                                                                                                   • MALDI-TOF mass spectrometer
                                                                               Dr. Dieter Stoll,   • Capillary electrophoresis
                                                                  Head of Bioanalytic/Synthesis
                                                                                                   • Various HPLC systems coupled to ms-,
                                                                                                     Ms/ms, UV-, diode array-UV-, fluores-
                                                                                                     cent- or light-dispersion detectors.
A. Jaworski at the MALDI-TOF mass spectrometer.




26   NMI Annual Report 2002/03
Peptide synthesis



Peptides are used in research and clinical       We synthesize:                              Contact:
medicine. We design and synthesize pep-          • Individual peptides, soluble peptides     Dr. Dieter Stoll
tides for over 50 research teams world             and peptide collections for epitope       phone: +49 7121 51530-811
wide. They appreciate not only the qual-           mapping, T-cell assays, as antigens for   stoll@nmi.de
ity, fast delivery and competitive pricing ,       antibody production, etc.
but also our ability to provide advice with      • Combinatorial peptide libraries           Mag. Pharm. Nils Clausen
respect to peptide design and applica-           • Modified peptides with:                   phone: +49 7121 51530-800
tion. While peptide-related R&D work is            -tags (FITC, TAMRA, Biotin...)            clausen@nmi.de
carried out at the NMI,                            -phospho-aminoacids
contract synthesis is performed at                 -non-proteinogenic aminoacids
the NMI Technology Transfer GmbH                   -lipid modification
(www.nmi-tt.de).                                   -cyclic and branched structures
                                                 • peptide conjugates
                                                 • peptide microarrays


                                                 All products can be obtained in quanti-
                                                 ties from 1 to 1000 mg and different
                                                 degree of purity.




                       High Throughput – automated peptidesynthesis.




                                                                                                        NMI Annual Report 2002/03   27
Biomedical technology



                                    Biomedical technology



                                    The medical technology industry is a
                                    very important economic factor for the
                                    south-western region of Germany. Tradi-
                                    tionally it was the domain of industries
                                    specialised in micromechanics, mechan-
                                    ics, optics, material sciences and textiles.
                                    More recently, life sciences and biotech-
                                    nology have opened new perspectives
                                    to medical technology, giving rise to the
                                    new field of biomedical technology. New
                                    medical products in which materials are
                                    combined with biomolecules, cells and
                                    tissues are now being developed.
                                    We are convinced that the interaction of
                                    biotechnology companies with the tra-
                                    ditional medical technology enterprises
                                    will give rise to new innovative products,
                                    thus sustaining economic growth in the
                                    region.
                                    The NMI, through its interdisciplinary
                                    structure, is playing an important role in
                                    the biomedical technology field, with a
                                    focus on
                                    • Neurotechnology
                                    • Tissue engineering
                                    • Development of medical products




                        Among the companies performing R&D with the
                        NMI in the area of neurotechnology, tissue engi-
                        neering and medical products we can mention:

                                    AESCULAP AG & Co. KG, Tuttlingen
                                    Altana Pharma AG, Konstanz
                                    Biopharm GmbH, Heidelberg
                                    CellMed AG, Alzenau
                                    DGF Stoess AG, Eberbach
                                    Labor Dr. Merk & Kollegen, Ochsenhausen
                                    Retina Implant AG, Tübingen
                                    TETEC AG, Reutlingen
Neurotechnology                               Tissue engineering                                         Development of medical
                                                                                                         products

The contact between the nerve cell            BioRegio Stern* placing a strong                           The task is: optimise the surface
and the microsystem is at the core of         emphasis on regenerative medicine
neuronal implants

Retina implants                               Cartilage regeneration                                     Improved functionality through
In cooperation with four research centres     Each year in Germany alone there                           surface coating
we are developing a retina implant to         are 900.000 new cases of arthrosis.                        The NMI develops coating procedures
restore vision in blind people (p. 31).       Autologous chondrocytes embedded in                        according to customer specifications
                                              biocompatible carrier materials represent                  (p. 37).
Biohybrid implants                            a new form of treatment (p. 33-34).
The vision is to integrate nerve cells with                                                              Qualification of cleanliness
microsystems into so-called biohybrid         Nerve regeneration                                         The handling, cleaning and sterilizing
implants. This could lead to new forms        Peripheral nerves can regenerate after                     of instruments, catheters and implants
of treatment of pain and paraplegia           injury. To facilitate and accelerate this                  poses considerable demands to the
(p. 32).                                      process, we develop hollow fibre nerve                     material of medical products. For this
                                              guides consisting of new biodegradable                     reason it is extremely important for the
                                              materials combined with Schwann cells                      manufacturers to apply validated meth-
                                              and nerve growth factors (p. 35).                          ods for the analysis of surfaces in order
                                                                                                         to ensure cleanliness and functionality
                                                                                                         of the product (p. 37).

                                              * BioRegio Stern: the acronym stern ist derived from
                                               the initials of the cities Stuttgart, Tübingen, Esslin-
                                               gen, Reutlingen and the region Neckar-Alb, which
                                               are collaborating in the field of tissue engineering.




          Neurotechnology
                                       Tissue engineering

Development of medical products

                                                                                                                        NMI Annual Report 2002/03    29
Biomedical technology
                                                        Neurotechnology




                                                        Neurotechnology



Contact:                                                Neurotechnological applications attempt        “A frequently quoted example for a
Dr. Alfred Stett                                        to treat sensory and motor impairments         successful neurotechnology application
phone: +49 71 21 5 15 30-70                             through selective electrical stimulation       is the cochlea implant for deaf patients.
stett@nmi.de                                            of nerves. For instance, clinical trials are   With this device after a learning period
                                                        evaluating the use of spinal, cortical or      hearing is partially restored. It is our
                                                        brainstem stimulation for the treatment        hope that in the next three years people
                                                        of chronic pain and Parkinson`s disease.       who turned blind because of retinitis
                                                                                                       pigmentosa will be treated with retina
                                                                                                       implants.”
                                                                                                                               Dr. Hugo Hämmerle,
                                                                                                                          Deputy managing director




                                                                                                       Implants for functional electrical stimula-
                                                                                                       tion are “active implantable medical
                                                                                                       devices” and must meet the require-
                                                                                                       ments of medical product legislation,
                                                                                                       such as biocompatibility, long-term
                                                                                                       stability, functionality, etc.




Prototype of an active, infrared-supplied implant. The cutout shows the
microelectrodes with the stimulation pixel, manufactured from titanium nitride.




 1 mm



                                                                                                                      Titannitrid Elektrode
                                                                                                                                50 x 50 µm




                                                                                                                         Stimulationspixel




                                                                                                          70 µm




30   NMI Annual Report 2002/03
Retina implants                                          room. A particular innovation resides in
Utopia or reality?                                       the electronic circuitry which drives each
                                                         individual electrode and allows trans-
Retina implants may restore vision in                    mission of contrasts in an illumination
patients with degenerative retina dis-                   range from 10 to 10.000 LUX (i.e. from
eases such as retinitis pigmentosa and                   moonlight to bright day values).
age-related macula degeneration. In                      The procedure for implanting the device
cooperation with the eye clinics of the                  directly under the retina has been
Universities of Tübingen and Regens-                     developed by the surgical team at the
burg, the Institute of Microelectronics                  eye clinic in Regensburg. The function-
Stuttgart (IMS) and other partners, we                   ality and biocompatibility of the retina
have developed an electronic implant                     implant will be tested in a clinical study
which will be placed under the retina.                   with eight patients in the year 2004.
The project was funded by the BMBF                       If successful, the device will be regis-
(FKZ 01KP0009).                                          tered (CE), produced and introduced to
The implant consists of a microphotodi-                  market by the Retina Implant AG, a new
ode array transforming incident light in                 enterprise founded by the partners who
electric signals which, upon electronic                  developed the implant.
amplification, should stimulate the optic
nerve and generate a pattern of phos-                    In 2003 the newly founded Retina
phenes which are processed in the visual                 Implant AG placed first in the inter-
cortex of the patient to yield an image.                 national business-plan competition of
With 1500 electrodes the implant should                  the state Baden-Württemberg and was
restore to a certain extent the ability of               awarded the “Gene Start Award”.
the patient to read and navigate in a

                                                                                                      Photography of the fundus of an eye with
                                                                                                      implanted prototype of retinal implant.




Assessment of the quality of vision obtainable with retinal implants. Depending on the quality of
coupling between the implant and the retina, the light-induced electrical stimulation gives rise
to the perception of separated „phosphenes“ which merge in the brain to generate shapes of
objects.




                                                                                                                        NMI Annual Report 2002/03   31
Biomedical technology
                                                      Neurotechnology




Contact:                                              Neuro implants
Prof. Dr. Burkhard Schloßhauer
phone: +49 71 21 5 15 30-20                           A new concept for the treatment               Neuron microprobe: a concept for
schlosshauer@nmi.de                                   of pain                                       coupling neurons with microdevices
                                                      Neurotechnology and tissue engineering        An implant has been developed
                                                      are combined to develop a new form            (Schlosshauer B. et al., 2001) which
                                                      of treatment of pain. The goal is an          consists of a biocompatible micro-
                                                      implantable system with genetically engi-     structure on which neuronal cells can
                                                      neered cells capable of producing and         grow and establish contact with muscle
                                                      releasing opioids under electrical stimula-   cells. Electric stimulation through this
                                                      tion. The system can be controlled by the     biohybrid implant induces contraction
                                                      patient. The cooperative project involves     of muscle fibres. This work provides a
                                                      the NMI, the International Neuroscience       “proof of concept” for the development
                                                      Institute (INI), the Foundation Neurobi-      of neuroprosthetic devices for paraplegic
                                                      onic and the CellMed AG. It is partially      patients.
                                                      funded by the BMBF (FKZ 0311962).




Biohybrid implant principle: spinal cord neurons
encapsulated in a containment extend neurites
through the sieve electrode implant which is
positioned onto the distal nerve stump. Electrical
stimulation of muscles is achieved via regenerating
neurites.




32   NMI Annual Report 2002/03
Tissue engineering




Tissue engineering



Biotechnology meets regenerative                        Regeneration of joint cartilage                      Contact:
medicine                                                A significant market is in the coming                Dr. Reinout Stoop
                                                                                                             phone: +49 7121 51530-805
Cells derived from a biopsy are grown                   Injury and wear of cartilage can cause               stoop@nmi.de
and differentiated in vitro, embedded                   severe pain and significant limitations of
in new biomaterials and re-implanted                    locomotion.
at the side of injury to promote tissue
regeneration. The NMI conducts R&D                      “Cartilage is a non-vascular non-inner-
with partners in basic research, clinics                vated tissue which cannot regenerate
and industry to elucidate the complex                   since chondrocytes are surrounded by a
interactions between cells and bioma-                   thick coat of matrix molecules. This was
terials. This work aims at the develop-                 the conventional opinion in the past. In
ment of new methods and products for                    today’s age of biotechnology, how-
regenerative medicine.                                  ever, we are on the best way of solving
                                                        this problem through the autologous
“We must learn how to induce tissue                     chondrocyte transplantation.”
remodelling in a sustainable fashion
                                                                           Dr. Jürgen Fritz, CEO, TETEC AG
through stimulation of transplanted
cells. An important step towards this                                                                        An SCID mouse model was established to investigate
goal, will be the development of                                                                             cartilage neosynthesis in a variety of biomaterials.
implants composed of new biomateri-                                                                          Human knee joint chondrocytes are isolated and
                                                                                                             seeded on a collagen membrane after an in vitro cul-
als capable of supporting the function
                                                                                                             tivation period. Eight weeks after implantation, the
of the embedded cells, so-called “smart                                                                      formation of a cartilage-like tissue can be observed
biomaterials.”                                                                                               in the center of the implant. This work is done in
                                                                                                             cooperation with the University of Tübingen, TETEC
         Dr. Reinout Stoop, Head of cell biology                                                             AG, and Aesculap AG & Co. KG and is supported in
                                                                                                             part by the BMBF (grant number 0312626).




Value Chain - regenerative procedures for tissue replacement.                                                                                        250 µm



          Biomaterials               Cell Systems                Tissue and Organ        Functionality,           Applications
                                                                 constitution            Testing


     plastics                   donor systems                   single cells         mechanical                 markets
     metals                     adult stem cells                mixed population     functionality              legislation
     textiles                   • cell production               organoids            secretory funct.           production
     membranes                  • identification                cell-material com-   structural funct.          ethical aspects
     microsystems               • progeny                       posites              animal models              clinical trials
     coatings                   • differentiation                                    compatibility
     structuring                • functional
                                • plasticity




                                                                                                                               NMI Annual Report 2002/03      33
Biomedical technology
                                                        Tissue engineering




Contact:                                                Autologous chondrocyte transplan-                       this reason, the company TETEC AG is
Dipl.-Ing. (FH) Karin Benz                              tation (ACT)                                            examining whether ATC in combination
phone: +49 71 21 5 15 30-827                            The autologous chondrocyte transplan-                   with new scaffolding biomaterials can
benz@nmi.de                                             tation (ACT) allows the formation of                    be used for the treatment of arthrosis of
                                                        new cartilage in the injured joint. In this             the knee in middle-aged patients. The
                                                        procedure a very small cartilage biopsy                 clinical study with NOVOCARD 3D is
                                                        is taken from the patient, the sample is                underway.
                                                        treated with specific enzymes to release                Further R&D work is now in progress at
                                                        the chondrocytes, these are expanded                    the NMI in cooperation with the com-
                                                        in vitro and then re-implanted into the                 panies TETEC AG, Aesculap AG & Co.
                                                        knee joint where they produce new car-                  KG and DGF Stoess AG. The research is
                                                        tilage tissue. During the past decade this              aimed at the development of “smart bio-
                                                        procedure has been applied with success                 materials” in combination with autolo-
                                                        to patients with knee joint injuries. It has            gous adult stem cells for bone/cartilage
                                                        not yet been adapted to the treatment                   regeneration.
                                                        of arthrosis, a disease with 900.000 new
                                                        cases each year in Germany alone. For

From the left: A. Scharstuhl, B. Gonser and R. Stoop.




                                                        High density cultures are an accepted method to
                                                        investigate the chondrogenic differentiation capacity
                                                        of adult mesenchymal stem cells. Cells are seeded
                                                        in u-bottom plates in a high density and a few days
                                                        later cell spheroids are formed. After several weeks
                                                        in culture the spheroids are analyzed for the expres-
                                                        sion of chondrogenic marker molecules. This project
                                                        is carried out with TETEC AG and the University of
                                                        Tübingen within the framework of the “Bioprofile“
                                                        programme “Regeneration Biology”
                                                        (FKZ 0312995).




                                                                                                                For the maintenance of the chondrocyte specific
                                                                                                                phenotype an optimal environment plays an impor-
                                                                                                                tant role. The Multiple Substrate Array (MSA™)(see
                                                                                                                page 20) developed by the NMI offers the possibility
                                                                                                                to test a diversity of extracellular matrix components
                                                                                                                for their effects on the chondrogenic phenotype. In
                                                                                                                this experiment porcine chondrocytes (nuclei in blue
                                                                                                                colour) are cultivated on the MSA™ and the synthe-
                                                                                                                sis of collagen type II, an important chondrocyte spe-
                                                                                                                cific marker molecule, is monitored (red: substrate
                                                                                                                spot consists of collagen type III).
                                                                                                                This work is done in cooperation with the Ortho-
                                                                                                                pedic Clinic of the University of Heidelberg, DGF
                                                                                                                Stoess AG, Altana Pharma AG, TETEC AG, Labor Dr.
                                                                                                                Koch & Kollegen with partial financial support by
                                                                                                                the “Landesstiftung Baden-Württemberg”
                                                                                                                (FKZ 4-4332.65-NMI/15).



34   NMI Annual Report 2002/03
Nerve regeneration                            Recombinant proteins                          Contact:
                                              Recombinant technology has been used          PD Dr. Hansjürgen Volkmer
Peripheral nerves can regenerate after        to produce proteins which promote neu-        phone: +49 7121 51530-44
injury. This process, however, is very        rite outgrowth. These growth-promoting        volkmer@nmi.de
slow and in many cases the regenerating       factors can be produced in considerable
nerves do not contact the tissue which        quantity in bioreactors. In combination       Prof. Dr. Burkhard Schloßhauer
they were innervating prior to injury. This   with the growth-factor GDF 5 they are         phone: +49 7121 51530-20
problem could be overcome through             embedded into biomaterials.                   schlosshauer@nmi.de
implantable biocompatible nerve guides.       Animal experiments in vivo show that
                                              these materials combined with vari-
Each year approximately 100.000               ous growth factors can promote nerve
patients with nerve injuries are treated      regeneration. This R&D work is carried
surgically and various types of nerve         out by the NMI in cooperation with the
guides are being employed, albeit with        Biozentrum and the Dept. of Neurology
only limited success. Significant progress    in Würzburg, and the enterprise Bio-
may be achieved, if nerve guides would        pharm GmbH, with partial funding from
contain specific nerve growth-promoting       the BMBF (FKZ 0311568 ).
factors and Schwann cells.
                                              Nerve guides containing Schwann
                                              cells
                                              Hollow fibres of biodegradable materi-
                                              als, e.g. polylactide, polyglycolide or
                                              collagen, can be seeded with Schwann
                                              cells and implanted at the site of nerve
                                              injury as guides for nerve regeneration.
                                              The aim of the project is to develop
                                              nerve guides which accelerate nerve
                                              regeneration, allow functional nerve          Neurites extend from neurons of a dorsal root gan-
                                                                                            glion positioned on a nerve guide fiber. Neurites are
                                              contacting of the target tissue and are       stained with a fluorescent label.
                                              degraded after a certain period of time
                                              (Schlosshauer, B. et al., 2003; Steuer, H.
                                              et al., 1999). This research is carried out
                                              in cooperation with the Deutsches Zen-
                                              trum für Biomaterialien und Organersatz
                                              e.V. (BMOZ), the ITV Denkendorf, the
                                              Berufsgenossenschaftliche Unfallklinik
                                              Tübingen and the University of Düssel-
                                              dorf, with partial funding from the BMBF
                                              (FKZ 01GN0111).




                                                                                                               NMI Annual Report 2002/03       35
Biomedical technology
                                                       Development of medical products




                                                       Development of medical products



Contact:                                               Technological advances ensure              share. For this reason the small and
Dr. Bernhard Schröder                                  economic growth                            medium enterprises form clusters with
phone: +49 71 21 5 15 30-52                                                                       researchers at academic and other
schroeder@nmi.de                                       The medical technology industry            institutions, with clinicians at university
                                                       represents a very important factor in      hospitals and with other enterprises
                                                       the economics of the south-western         in order to establish new technologies
                                                       region of Germany. Surgical instruments,   which can later be used in the develop-
                                                       catheters, implants and other medical      ment of innovative medical products.
                                                       products undergo considerable wear
                                                       through use, cleaning and sterilisation.   The NMI has expertise relevant to the
                                                       The materials, in particular the surface   medical product industry in the follow-
                                                       of the medical product, must meet the      ing areas:
                                                       highest standards with respect to long     • Surface modification through passiva-
                                                       term stability, functionality and clean-     tion and coatings (p.45)
                                                       liness.                                    • Qualification of the cleanliness of
The NMI is partner of the „Kompetenzzentrums
Minimal Invasive Medizin + Technik Tübingen-Tuttlin-                                                medical products at the end of the
gen“, an R&D alliance between enterprises, research    Innovative enterprises in the field of       production process and after cleaning
institutions and clinics. New medical products are     medical technology must invest large         for reuse
beeing developed with support of a grant from the
State Baden-Württemberg (FKZ 4-4331.1/67)
                                                       sums of money for R&D in order to          • Improvement of the material properties
                                                       maintain and increase their market-          of medical products in order to extend
                                                                                                    their life-time




Some of the companies which we assist in their product development are:
ADELHELM LubriCoat GmbH, Eningen                       Geister Medizintechnik GmbH, Tuttlingen    MTI Technische Instrumente GmbH,
AESCULAP AG & Co. KG, Tuttlingen                       Georg Leibinger Apparatebau, Mühlheim      Tuttlingen
ALCON PHARMA GmbH, Freiburg                            HAMO AG, Pieterlen/Schweiz                 Ohst Medizintechnik AG, Rathenow
ALUMEDICA GmbH, Geisingen                              Heine Optotechnik GmbH & Co. KG,           PI Precision Implants AG, Aarau/Schweiz
APPLIEDSENSOR GmbH, Reutlingen                         Herrsching                                 Plasma Ionic GmbH, Besigheim
Berchtold GmbH & Co. KG, Tuttlingen                    Heinz Kurz GmbH, Dußlingen                 Polyzenix GmbH, Ulm
Biedermann Motech GmbH,                                HENKE-SASS, WOLF GmbH, Tuttlingen          PRECISWISS AG, Hünenberg/Schweiz
VS-Schwenningen                                        IIP Technologies GmbH, Bonn                QualiMed Innovative Medizinprodukte
BOEHRINGER INGELHEIM PHARMA KG,                        IMPLANTCAST GmbH, Buxtehude                GmbH, Winsen
Biberach                                               Institut für Med. Mikrobiologie und        REMED GmbH, Friedeburg
BORER CHEMIE AG, Zuchwil/Schweiz                       Krankenhaushygiene, Tübingen               RICHARD WOLF GmbH, Knittlingen
BSL BIOSERVICE Scientific Laboratories                 JAKOUBEK Medizintechnik GmbH,              Roche Diagnostic GmbH, Mannheim
GmbH, Planegg                                          Liptingen                                  Sektion für Minimal Invasive Chirurgie
Christoph Miethke GmbH & Co. KG,                       Kapp GmbH, Dürbheim                        der Universität Tübingen, Tübingen
Kielnmachnow                                           KARL STORZ GmbH & Co., Tuttlingen Ker-     Smith & Nephew Orthopaedics GmbH,
CRANIUM TELEMETRICS GmbH, Hannover                     amed Medizintechnik GmbH, Mörsdorf         Tuttlingen
Dentaurum J.P. Winkelstroeter KG, Ispringen            Kreidler Medizin-Technik GmbH,             SMP GmbH, Tübingen
Disetronic Medical Systems AG,                         Wurmlingen                                 SQ Products AG, Steinhausen/Schweiz
Burgdorf/Schweiz                                       Mathys Medizintechnik AG, Bettlach         STERIPAC GmbH, Calw
EBRO ELECTRONIC GmbH & Co. KG,                         medical & precision technology GmbH,       Stratec Medical, Oberdorf/Schweiz
Ingolstadt                                             Annaberg-Buchholz                          STRYKER TRAUMA AG, Selzach/Schweiz
EPFLEX Feinwerktechnik GmbH, Dettingen                 MESOTEC Gesellschaft für medizinische      Surface Contacts GmbH, Saarbrücken
ESKA Implants GmbH & Co., Lübeck                       Sensortechnik mbH, Hannover                SURGICAL INNOVATIONS Ltd., Leeds/
ETHICON ENDO-SURGERY, INC.,                            METRAX GmbH, Rottweil                      Großbritannien
Cincinnati/USA                                         MINITUBES ZAC Technisud, Grenoble/         TECHNO-COAT Oberflächentechnik GmbH,
EUROflex G. Rau GmbH, Pforzheim                        Frankreich                                 Zittau
Forestadent Bernhard Förster GmbH,                     MITT Kompetenzzentrum Minimal Invasive     TONTARRA Medizintechnik GmbH,
Pforzheim                                              Medizin + Technik Tübingen-Tuttlingen,     Wurmlingen
FRÜH VERPACKUNGEN AG, Schwerzenbach/                   Tübingen                                   TRANSLUMINA GmbH, Hechingen
Schweiz                                                                                           Ulrich GmbH & Co. KG Medizintechnik,
                                                                                                  Ulm-Jungingen


36   NMI Annual Report 2002/03
Improvement of surface properties                    Qualification of cleanliness                     Contact:
Functional improvement through coating               What is clean?                                   Dr. Rudolf Reichl
                                                                                                      phone: +49 7121 51530-57
In order to modify the surface proper-               It goes without saying that cleanliness          reichl@nmi.de
ties of a medical product (hydrophilic/              and hygiene are basic concepts for
hydrophobic, hardness, tribological                  manufacturers and users of medical
qualities, etc.) the NMI develops specific           products. Surgical instruments, endo-
procedures to coat medical devices                   scopes, catheters, etc. must meet the
with diamond-like carbon (DLC), with                 highest functional and hygienic stand-
polymers (e.g. polyzene). Electrophoretic            ards. A simple solution is to manufacture
coating, galvanic coating (hard gold for             and employ only disposable products.
endoscopes), electropolishing, etc. are              However, in view of the ever increasing
some of the methods we employ.                       cost of medical care, there is a strong
                                                     demand for reusable products. This is
“We develop coating procedures accord-               only possible if the effectiveness of the
ing to the specifications of each cus-               cleaning and sterilisation procedures can
tomer. The products are tested to assure             be demonstrated.
the quality of the coating with respect to
its adhesion and stability under mechani-            “An objective assessment of the cleanli-
cal loading, ultrastructure, chemical                ness of a surface was not possible so
composition, etc. The validated coating              far. We have defined objective criteria          R. Reichl and B. Schröder inspect a catheter.
procedure is transferred to the client. “            and have validated the procedures to
                                                     be employed in the cleaning of reusable
                         Dr. Bernhard Schröder,
                     Head of surface technology      medical products.”
                                                                                 Dr. Rudolf Reichl,
                                                                        Head of interface analysis



                                                     To determine the cleanliness of a sur-
                                                     face, scientists at the NMI apply electron
                                                     microscopy and other sophisticated spec-
                                                     troscopy methods. All relevant tests and
                                                     analytical procedures were developed
                                                     and validated in a project funded by the
                                                     EU (FKZ G6ST-CT-2001-50121).

                                                     Next to the evaluation of the degree
                                                     of cleanliness of a surface of a medical
                                                     product processed for re-use, it is equally
                                                     important to verify that the functional-
Nitinol stent with Diamond Like Carbon (DLC) coat-   ity of the product is still intact. To this
ing. The layer generates a typical rainbow effect.   goal, the NMI has established a series of
                                                     product-specific tests to examine mate-
                                                     rial qualities such as hardness, friction,
                                                     colour, corrosion and mechanical char-
                                                     acteristics under load. These tests allow
                                                     to monitor processes such as cleaning
                                                     and sterilisation or passivation, and are
                                                     equally well suited for damage analysis
                                                     of medical products.




                                                                                                                         NMI Annual Report 2002/03    37
Surface and interface technology



                                       Surface and interface technology



                                       Our competence in surface and inter-
                                       face technology results from our many
                                       years of research of those physical and
                                       chemical processes which are essential to
                                       the production of layers, of micro- and
                                       nanostructures, as well as to the analysis
                                       of materials, surfaces and interfaces.
                                       Our know-how in this field finds its
                                       application not only in material sciences,
                                       but also in projects in biotechnology
                                       (p. 21), neurotechnology (p. 30) and
                                       biomedical technology (p. 36).

                                       Surface, interface and microstructure
                                       analysis are essential for the develop-
                                       ment and validation of quality control
                                       standards and for damage analysis
                                       of materials. This type of analysis can
                                       provide insight into the molecular proc-
                                       esses which take place at interfaces and
                                       is therefore used in R&D in the material
                                       sciences. For example, in the field of tri-
                                       bology there exists a large empiric know-
                                       how concerning the interaction under
                                       friction of materials and lubricants. The
                                       molecular mechanisms of these proc-
                                       esses, however, are poorly understood.
                                       For this reason the NMI is involved in the
                                       analysis of micro- and nanostructures of
                                       the interface of tribological systems.

                                       In collaboration with competent indus-
                                       trial and academic partners we work in
                                       the following areas:

                                       •   Microsystems and nanotechnology
                                       •   Functional surfaces
                                       •   Bonding, adhesion- and tribo-
                                           logical systems




                          We collaborate with the following enterprises
                          and organisations:
                                       ADA Abgaszentrum der
                                       Automobilindustrie, Weissach
                                       BAL-TEC, Balzers/Lichtenstein
                                       Cytocentrics CCS GmbH, Reutlingen
                                       Evotec Technologies GmbH, Hamburg
                                       FVA Forschungsvereinigung
                                       Antriebstechnik e.V., Frankfurt
                                       Greiner Bio One AG, Frickenhausen
                                       Kleindiek Nanotechnik, Reutlingen
Microsystems and nano-                        Functional surfaces and                        Bonding, adhesion and tribo-
technology                                    interfaces                                     logical systems

Motors for novel applications                 The world of microstructures                   Processes at the interfaces of
                                                                                             materials

Sensor arrays and multiwells with             Interface and microstructure                   Bonding and adhesion
microelectrodes                               analysis                                       New technologies are in demand to join
Sensor arrays for electrical stimulation      State-of-the-art microscopy and spec-          components of medical products (p. 47).
and recording of cells and tissues in vitro   troscopy is employed to analyse the
and in vivo find their application in drug    ultrastructure of surfaces. The informa-       Tribological systems
discovery, biotechnology and neuro-           tion is of great value to our customers        Tribology is the discipline which studies
technology. The production of stable          in order to optimise the quality of their      the interactions between two surfaces
and biocompatible devices for electrical      products (p. 44).                              moving against each other (p. 48).
stimulation represents a considerable
technological challenge (p. 40).              Functional surfaces
                                              Our clients request that we improve
BioMEMS                                       the surface qualities of their products:
A growing market in the field of              e.g. increase the resistance to wear
microsystem technology is the integra-        and corrosion, prevent certain reactions
tion of microelectronic and mechanical        between the surface of the product and
systems (BioMEMS) for applications in         components of its environment, render
life sciences and medicine. A system for      a surface biocompatible for biomedical
the separation, accumulation and analy-       applications, etc. (p. 45).
sis of biological micro- and nanoparticles
has been developed (p. 41).

Nanoworkbench
The name stands for rapid prototyp-
ing of 3-D-nanostructures and repairs
in nanoscale inside a scanning electron
microscope (p. 42).

Focused ion beam (FIB) electron
microscope and workstation in one
instrument
In cooperation with the company LEO
we have developed a new CrossBeam-
Workbench which opens new perspec-
                                                      “God created the volume,
tives in micro- and nanotechnology
(p. 43).
                                                  the devil the surface”
                                                                                          Wolfgang Pauli, Nobelpreis Physik 1945




LEO Elektronenmikroskopie GmbH,
Oberkochen
MAN, Nürnberg
Mediagnost GmbH, Reutlingen
Multi Channel Systems MCS GmbH,
Reutlingen
Robert Bosch AG, Stuttgart
ZF Friedrichshafen AG, Friedrichshafen


                                                                                                            NMI Annual Report 2002/03    39
Surface and interface technology
                                                          Microsystems and nanotechnology




                                                          Microsystems and nanotechnology



Contact:                                                  Motors for innovation                              Sensor arrays and multiwells with
Dr. Wilfried Nisch                                                                                           microelectrodes
phone: +49 71 21 5 15 30-40
nisch@nmi.de                                              During the past decade microsystem                 Microelectrode arrays (MEAs)
                                                          technology has been applied to produce             Microelectrode arrays find their applica-
                                                          components for innovative products                 tion in drug discovery, in biotechnology
                                                          such as medical devices, robots, instru-           (p.21-22) and in medical devices (e.g.
                                                          ments for measurement and control of               implants for muscle and nerve stimula-
                                                          automated processes, scientific instru-            tion). Not only must such devices fulfill
                                                          ments. Microsystems are also employed              specific technical criteria, but they must
                                                          in optics, electronics and manufactur-             also be biocompatible and exhibit long-
                                                          ing system engineering. At present the             term stability.
                                                          world-wide market for products based               At the NMI we employ a variety of sput-
                                                          on microsystem technologies is about               ter procedures to generate microelec-
                                                          50 bn $.                                           trode arrays with specific properties. For
                                                          Nanotechnology is seen as a new key-               instance, we produce titanium or iridium
                                                          technology of the next decade. There               microelectrodes with a large inner
                                                          is, however, a large gap between vision            surface, such that a high charge can be
                                                          and reality. Germany is strong in the              transmitted to tissues or cells, without
                                                          nanosciences, but week in bringing this            concomitant electrode polarisation or
                                                          scientific knowledge to market.                    electrochemical reactions at the interface
                                                                                                             between cell and electrode. For the com-
                                                          “The NMI develops micro- and nano-                 pany Multi Channel Systems MCS GmbH
                                                          systems for medical products. Some of              we produce high density microelectrode
                                                          these have already entered the market.             arrays with 264 electrodes on glass
Flex MEA: with a micromanipulator a polyimide
                                                          Our strength is the ability to handle a            substrate (HD MEA), flexible polyimide
based flexible MEA is placed above the electrodes of
a conventionell glass MEA in order to simultaneously      variety of materials which are bio-com-            microelectrode arrays (flex MEA) and
stimulate and record brain slices.                        patible and bio-stable”.                           MEAs with suction wholes to permit
                                                                                                             tissue adhesion.
                                                                                           Wilfried Nisch,
                                                                            Head of Physical Technologies
                                                                                                             Thin-layer and microstructure
                                                                                                             technology
                                                                                                             The NMI has all relevant equipment and
                                                                                                             the adequate infrastructure to produce
                                                                                                             custom-specific micro- and nanostruc-
                                                                                                             tured products.
                                                                                                             • Clean room
                                                                                                             • Plasma vapour deposition
                                                                                                             • Sputter
                                                                                                             • Plasma enhanced chemical vapour
                                                                                                               deposition
                                                                                                             • Plasma etching
                                                                                                             • Chemical etching
                                                                                                             • Photolitography (2-6 inches)




                                                       HD-MEA: microelectode array with 256 electrodes
                                                       integrated in a microelectronic chip package


40   NMI Annual Report 2002/03
Multiwells with electrodes                    Bio MEMS                                       Contact:
Microtitre plates with 96, 384 and 1536       Manipulation of smallest particles             Dr. Martin Stelzle
wells have become standard laboratory                                                        phone: +49 7121 51530-75
equipment. Pipetting robots for the           There is a rapidly growing market for          stelzle@nmi.de
precise dispensing of cells and reagents      the application of microelectronic and
into such multiwell plates combined with      mechanical systems in biological sciences
appropriate readout systems are used          (bio MEMS). With partial support of the
in drug discovery by the pharmaceuti-         BMBF (FKZ 13N7939) and in coopera-
cal industry for rapid screening of the       tion with the Fraunhofer Institute for
effects of compounds on cell function.        Biomedical Technology, St.Ingbert, the
In some instances one would like to           Humbold University in Berlin and the
measure the effects of compounds on           companies Evotec Technologies GmbH
the electric potentials of cells (p.22). To   and Mediagnost GmbH, we have                   Prototype of the NanoVirDetect system with fluidic
                                                                                             ports, microchannels and 3-D-electrode-struc-
this goal we develop microtitre plates        developed a system for the separation
                                                                                             tures for separation, accumulation and trapping
with wells equipped with microelec-           and analysis of cells, cell organelles,        of particles in solutions of biological probes using
trodes. When electrogenic cells such as       bacteria and viruses. The microsystem          dielectrophoretic forces. The system unifies fluidics
neurons or heart cells are dispensed into     contains micrelectrodes and channels           and electrical contacting on a chip and fits in a self-
                                                                                             adjusting device.
these wells, suction channels effectuate      for microfluidics. Particles in a mixture
the adhesion of the cells to the elec-        can be separated according to size by
trode. This allows the electrophysiologi-     subjecting the mixture to electrical fields.
cal analysis of the effects of compounds      Separation of particles is a function
on cells. This project is partially funded    of their respective dielectric constant ,
by the BMBF (FKZ 16SV1744) and is             frequency, shape and forces influencing
carried out in collaboration with the         particle flow through the microchan-
University of Freiburg and the companies      nels. Thus, the development of a device
Multi Channel Systems MCS GmbH and            which allows separation, enrichment and
Greiner Bio One AG.                           recovery of particles which can then be
                                              subjected to further analysis, represents
                                              a first step towards the establishment of
                                              a “lab-on-the –chip” system (Dürr, M. et
                                              al. 2001; Kentsch, J. et al. 2003).




                                                                                               From the left: W. Nisch, A. Stett and M. Stelzle.




                                                                                                                 NMI Annual Report 2002/03         41
Surface and interface technology
                                                  Microsystems and nanotechnology




Contact:                                          Nano-Workbench                                         This nano-workbench was developed
Dr. Claus Burkhardt                               A tool for the nano-world                              with partial support from the BMBF (FKZ
phone: +49 71 21 5 15 30-55                                                                              13N8004) in cooperation with the Uni-
burkhardt@nmi.de                                  Rapid prototyping and repair of 3-D-                   versity of Tübingen and the companies
                                                  nanostructures can only be performed                   Kleindiek Nanotechnik, LEO Elektronen-
                                                  under observation in an electron                       mikroskopie GmbH and BAL-TEC GmbH
                                                  microscope. The so-called nano-work-                   (Burkhardt, C. et al. 2002; Bauerdick,
                                                  bench allows construction of 20-50                     S. et al. 2003).
                                                  nanometer structures in a high-resolu-
                                                  tion field-emission scanning electron-
                                                  microscope by means of electron beam
                                                  litography, additive nanolitography and
                                                  ion etching or through electron beam
                                                  induced material deposition from the
                                                  gas phase. The nanostructures can be
                                                  manipulated through microlevers driven
                                                  by a nanomotor (e.g. for transmission
                                                  electron microscope analysis).

Two micromanipulators within the chamber of the
nano-workbench.

                                                                                                         3D structures built by electron beam-CVB (EB-CVD)




                                                  Gemini-NanoWorkbench
                                                  A combination of high resolution field emission scanning electron microscope
                                                  with gas injection for EB-CVB, micromanipulators and ion guns.




42   NMI Annual Report 2002/03
Focused ion beam (FIB)                                                        „Nano Violinist“ (Design Tarek Lutz,
                                                                              www.tarek-lutz.de)
Electron microscope and workstation
at once

In cooperation with the company LEO
Electronen Microscopie GmbH we have
developed a new procedure which finds
its application in the CrossBeam®-work-
station. It has always been a dream of
material scientists to be able to see the
micro- and nanostructures of a surface
in a scanning electron microscope and
at the same time be able to manipulate
and process the sample for further analy-
sis. This vision is now reality, thanks to
the new CrossBeam®-workstation.              Furthermore, the workstation allows to
Within the scanning electron microscope      build 3-D-micro- and nanostructures by
the sample can be milled and cut by          means of ion beam induced material
means of a focused ion beam and the          deposition (p.42).
newly exposed surfaces can be visualized
by ion or electron optics. We have used      “With the CrossBeam®-workstation we
this CrossBeam®-workstation to identify      have a universal instrument for probe
at the nanostructural level the cause        analysis- and preparation. This technol-
of functional defects of semiconductor       ogy opens new avenues for material
chips, defects in the layers of coated       sciences.”
surfaces, the type of damage caused
                                                   Dr. Claus Burkhardt, Physical technologies
by corrosion, by friction in tribologic
systems or by metal fatigue.




                                                                                                  Examples for applications of the CrossBeam®
                                                                                                  workstation:
                                                                                                  A Cross section of a semiconductor device
                                                                                                  B Rapid prototyping of nano structures:
                                                                                                    micro pipette in glas




    2 µm                                         2 µm

A                                            B



                                                                                                                     NMI Annual Report 2002/03   43
Surface and interface technology
                                                      Functional surfaces and layers




                                                      Functional surfaces and layers



Contact:                                              Coronary stents with an antithrombo-         Interface and microstructure
Dipl.-Phys. Werner Dreher                             genic surface, biostable passivation of a    analytics
phone: +49 71 21 5 15 30-59                           neuro-implant, biocompatible electrodes,     A detailed understanding of the proc-
dreher@nmi.de                                         insulating layers on BioChips, control       esses which influence and alter the
                                                      of wetting in a microdispensing system,      surface of materials during product
                                                      suppression of protein adsorption in         manufacturing or use is of primary
                                                      microfluidic systems, coating of surfaces    importance to the producer. The NMI
                                                      to promote cell adhesion... these are just   has the most modern equipment and
                                                      examples for the many projects in which      uses validated procedures to analyze the
                                                      the NMI know-how in surface- and             physico-chemical properties of surfaces,
                                                      interface technology finds its practical     down to the atomic level.
                                                      application.
                                                      A common theme is the surface modifi-        “The smaller the structure, the more
                                                      cation in order to achieve a specific cell   involved is the procedure we need to
                                                      or tissue interaction with the material.     apply, in order to obtain valid informa-
                                                                                                   tion. One of our strengths is the ability
                                                                                                   to prepare probes for microstructure
                                                                                                   analysis from all kinds of materials.”

                                                                                                                Werner Dreher, Head of interface
                                                                                                                    and microstructure analytics




From the right: W. Dreher, P. Beckmann and N. Kern.




44   NMI Annual Report 2002/03
Layers                                         An extensive and modern instrumen-          Contact:
The need for interdisciplinary                 tation is available to the NMI to per-      Dr. Bernhard Schröder
cooperation                                    form spectroscopic and microscopic          phone: +49 7121 51530-52
                                               analyses                                    schroeder@nmi.de
A core competence of the NMI is the            • Surface topography and dimensional
modification and functionalisation of            accuracy is analyzed by:
surfaces for medical technology and bio-         light microscopy, scanning electron
technology products. The development             microscopy (REM, ESEM), profilometry,
of functional layers requires an intense         atomic force microscopy (AFM), confo-
cooperation between experts in biologi-          cal laser scanning microscopy (CLSM),
cal and material sciences.                       transmission electron microscopy
                                                 (TEM), scanning tunnel microscopy
Surface modification with diamond-               (STM).
like-carbon (DLC)                              • Surface chemistry and atomic compo-
Surface modifications through DLC find           sition is analyzed by:
their application in a variety of fields,        Photoelectron spectroscopy (XPS),
such as medical technology, engine con-          secondary ion and secondary neutral
struction and textile industry (needles).        particle mass spectroscopy (SIMS,
The DLC layer creates a hard, yet elastic,       SNMS), Raman spectroscopy, optical
surface which is abrasion and corrosion          spectroscopy (UV/VIS/IR) and Fourier
resistant, chemically inert, has particu-        transformed infrared (FTIR) spectros-
larly good tribologic properties, is biosta-     copy, electron energy loss spectroscopy
ble, biocompatible, antithrombogenic             with transmission electron microscopy
(stents) and prevents leakage of atoms           (EELS/TEM), energy dispersive x-ray
from the product material into body              (EDX/TEM), EDX environmental scan-
fluids (e.g. prevention of allergy caused        ning electron microscope (EDX ESEM),
by leakage of nickel from implants).             x-ray fluorescent analysis (XRFA).
In our plasma enhanced chemical vapor          • Inner solid state properties are
deposition (PECVD) plant we can adjust           analyzed by:
the process parameters such that also            Cross section analysis (SEM, EDX)
objects of complex geometry can be               under visual control in the Cross-
uniformly coated. Furthermore, specific          Beam®-workbench (p.43).
coating characteristics such as thermo-
stability, adhesive properties, color and      Some of the above analytical methods
electric conductance can be influenced         are performed in accredited labora-
in order to obtain a product which meets       tories according to DIN EN ISO/IEC
the needs of the customer.                     17025. The QM system is applied to all
We develop custom-specific coating             methods, regardless of the accreditation
processes which are then transferred to        status.
the client. Alternatively, we can perform
serial coating of products in-house.




                                                                                                      NMI Annual Report 2002/03   45
Surface and interface technology
                                                     Functional surfaces and layers




Contact:                                             Passivation of stainless steel                    Protective biostable and biocom-
Passivation                                          In the field of medical technology, phar-         patible coatings for neuroprosthesis
Dr. Rudolf Reichl                                    maceuticals and food processing many              Silicon oxide becomes corroded in body
phone: +49 71 21 5 15 30-57                          machines and instruments are made of              fluids (Hämmerle, H. et al. 2002). The
reichl@nmi.de                                        stainless steel. Cleaning, disinfection and       NMI has developed biocompatible and
                                                     sterilization, however, can cause consid-         microstructured coatings which protect
Functional surfaces                                  erable wear and damage to the stainless           the silicon for at least one year.
Dr. Martin Stelzle                                   steel surface. To protect the material
phone: +49 71 21 5 15 30-75                          from corrosion, the steel surface is              Controlled wetting in microfluidic
stelzle@nmi.de                                       enriched with chromium and chromium               sytems
                                                     oxides, a process called “passivation”.           Covalent functional modifications of the
                                                     At the NMI we apply interface analysis            surface of components of microfluidic
                                                     (XPS and REM) to verify the quality of            systems influence the flow properties.
                                                     passivation.

                                                     “In a cooperative project with the MITT
                                                     Kompetenzzentrum Minimalinvasive
                                                     Medizin+Technik Tübingen-Tuttlingen
                                                     and with 17 enterprises ranging from
                                                     steel producers to end-users, we
                                                     have established analytical procedures
                                                     to ensure the quality of passivated
                                                     surfaces.”

                                                                                  Dr. Rudolf Reichl,
                                                              Interface and microstructure analytics
Dispensing of nanolite-droplets by a TopSpot
dispenser with selectively coated outer surface.
The coating prevents intermixing of solutions from
different outlets and ensures long-term stable and
well-defined dispensing properties of the device.




                            Some of the companies which we assist in product development are:
                            Adolf Schweickhardt GmbH & Co. KG,               Henke-Sass, Wolf GmbH, Tuttlingen
                            Tuttlingen                                       Jakoubek Medizintechnik GmbH, Liptingen
                            Aesculap AG & Co. KG, Tuttlingen                 Karl Storz GmbH & Co., Tuttlingen
                            Boner Galvanotechnik, VS-Schwenningen            MKK GmbH, Oldenburg
                            Borer Chemie AG, ZUCHWIL                         nopa instruments Medizintechnik GmbH,
                            Brasseler GmbH & Co. KG, Lemgo                   Tuttlingen
                            Eisold GmbH, Balgheim                            Richard Wolf GmbH, Knittlingen
                            EPflex Feinwerktechnik GmbH,                     Rudolf Medizintechnik GmbH & Co. KG,
                            Dettingen/Erms                                   Fridingen/Do.
                            Forestadent Bernhard Förster GmbH,               ThyssenKrupp Nirosta GmbH, Krefeld
                            Pforzheim                                        Tontarra Medizintechnik GmbH, Wurmlingen
                            Geister Medizintechnik GmbH, Tuttlingen          Translumina, Hechingen




46   NMI Annual Report 2002/03
Bonding, adhesion and tribologic systems




Bonding, adhesion and tribologic systems



Physico-chemical processes at the            Bonding                                       Contact:
interface of two materials govern the        Although a very ancient technology,           Dr. Bernhard Schröder
bonding, adhesive and tribologic proper-     bonding is much less frequently used          phone: +49 7121 51530-52
ties of interacting bodies. So far empiric   then welding or soldering. This is surpris-   schroeder@nmi.de
approaches were at the basis of techni-      ing, since almost all materials can be
cal solutions in these areas. The NMI is     bonded under conditions which do not
involved in studies aiming to provide a      alter the material characteristics of the
theoretic foundation which should allow      bonded components.
to describe and anticipate the effects of    The NMI has expertise in the systematic
various parameters on bonding, adhe-         selection of bonding systems, surface
sion and friction. This will allow a more    preparation for bonding, examination of
rational approach for the manufacturing      bonding strength and other parameters.
of medical products, engineering of drive    To better meet the specific needs of
systems and components, chemical com-        our clients, we cooperate with several
position of adhesive and lubricants, etc..   institutes and manufacturers of bonding
                                             materials (Brockmann,W. et al. 2004).
System development in these areas is
very empirical. Numerous small and
medium enterprises are interested in
practical solutions which can be applied
to their specific product manufacturing
process. The FVA Forschungsvereinigung
Antriebstechnik (Research association
for drive engineering) is an association
of many enterprises with which the
NMI collaborates very intensively. Other
collaborations take place with various
academic institutions, the Institute of
Tribology (FH, Mannheim), the Institute
for materials for electrotechnology (Uni-
versity of Ulm) and the Research center
for cog wheel and gear construction
(Technical University of Munich).




                      Some of the companies which we assist in product development are:
                      Chemie Technik GmbH, Vöhringen                KaVo Dental GmbH & Co. KG, Biberach
                      DaimlerChrysler AG, Ulm                       KHS Steiert, Simonswald
                      ebm-papst St. Georgen GmbH & Co. KG,          Maxon Motor GmbH, Sexau
                      St. Georgen                                   Robert Bosch GmbH, Gerlingen
                      FAG Kugelfischer AG, Schweinfurt              Dr. Tillwich GmbH Werner Stehr, Horb
                      Follmann & Co., Minden                        Uzin Utz AG, Ulm
                      Fuchs Petrolub AG, Mannheim                   Zahnradfabrik Friedrichshafen AG,
                      FVA Forschungsvereinigung                     Friedrichshafen
                      Antriebstechnik e.V., Frankfurt




                                                                                                           NMI Annual Report 2002/03   47
Surface and interface technology
                                       Bonding, adhesion and tribologic systems




Contact:                               Tribologic systems                            been subjected to prolonged load under
Dr. Peter Beckmann                                                                   various conditions. The results provide
phone: +49 71 21 5 15 30-51            Tribology is the science which examines       the first insight into the molecular events
beckmann@nmi.de                        the reactions which take places at the        leading to material fatigue and damage.
                                       interface of two bodies moving against
Dr. Walter Holweger                    each other under load. It examines fric-      Damage to gears and motor
phone: +49 71 21 5 15 30-863           tion, wear and lubrication. The physico-      components
holweger@nmi.de                        chemical reactions of this process are        The State of Baden-Württemberg has
                                       poorly understood. The NMI examines           provided funding (AZ 4-4332.62-NMI/
                                       the interactions between the lubricants       16) for a research project aimed to
                                       (different oils with various additives) and   develop a system for the early recogni-
                                       the surfaces of the moving bodies under       tion of damage to gears and cog wheels.
                                       load by means of chemical, interface and      The system should substitute the com-
                                       microstructure analysis.                      monly used long-term wear assays. Eight
                                                                                     enterprises are participating in the study.
                                       Tribologic layer                              Both the ultrastructure of the material
                                       In a research project (FVA 289) with the      (p. 43) and the chemical composition of
                                       Forschungsvereinigung Antriebstechnik         the lubricant are at the center of these
                                       (Research association for drive engineer-     investigations.
                                       ing) and the Research center for cog
                                       wheel and gear construction of the            Micro-gears
                                       Technical University of Munich, we have       Miniaturized motors and micro-gears
                                       examined at the ultrastructural level, i.e.   are essential components of numerous
                                       in a depth of 2 µm , the surface of gears     high-technology devices, such as drills
                                       and anti-friction bearings which had          for dental instruments of the company
                                                                                     KaVo Dental GmbH Co. KG, or parts
                                                                                     for injection molding produced by the
                                                                                     company KHS Steinert, or ceramic
                                                                                     components of very small aggregates
                                                                                     produced by Maxon Motor GmbH. With
                                                                                     these partners and with partial support
                                                                                     by the BMBF (FKZ 16SV1644) we analyze
                         Gear wheels                                                 the tribological parameters in order to
                                                                                     develop better products.




48   NMI Annual Report 2002/03
Staff




Total staff                           Scientists and engineers
(12/31/03)
                                      Dr. Brigitte Angres, biologist
35   scientists                       Dr. Peter Beckmann, physicist
 8   Ph.D.students                    Karin Benz, engineer
 2   undergraduates                   Stefanie Breisch, engineer
 5   trainees                         Dr. Claus Burkhardt, physicist
10   engineers                        Nils Clausen, pharmacist
13   technicians                      Werner Dreher, physicist
 7   assistants                       Dr. Cornelia Fricke, biologist
 7   administration and secretariat   Prof. Dr. Elke Guenther, biologist
                                      Dr. Nadja Gugeler, biologist
                                      Dr. Hugo Hämmerle, biologist
                                      Michael Hartmann, engineer
                                      Thoralf Herrmann, physicist
                                      Bernd Holder, engineer
                                      Dr. Otto Inacker, chemist
                                      Dr. Andreas Jaworski, nutritionist
                                      Dr. Thomas Joos, biochemist
                                      Andrea Karl, engineer
                                      Karin Klaß, engineer
                                      Dr. Conny Kuschel, biologist
                                      Dieter Martin, engineer
                                      Enzio Müller, Ph.D., biologist/
                                      biochemist
                                      Dr. Marc Niere, biologist
                                      Dr. Wilfried Nisch, physicist
                                      Dr. Ulrich Pehl, biologist
                                      Michael Quellmalz, engineer
                                      Dr. Rudolf Reichl, physicist
                                      Dr. Uta Rheinweiler, biologist
                                      Ingrid Ruchay, engineer
                                      Ralf Rudorf, engineer
                                      Dr. Alwin Scharstuhl, biologist
                                      Prof. Dr. Burkhard Schloßhauer,
                                      biochemist
                                      Monika Schrenk, chemist
                                      Dr. Bernhard Schröder, chemist
                                      Dr. Bernhard Stein, physicist
                                      Dr. Martin Stelzle, physicist
                                      Dr. Alfred Stett, physicist
                                      Heiko Steuer, biologist
                                      Dr. Dieter Stoll, chemist
                                      Dr. Reinout Stoop, biologist
                                      Dr. Sabine Sturany, biologist
                                      Dr. Markus Templin, biologist
                                      PD Dr. Hansjürgen Volkmer, biologist
                                      Dr. Frank Weise, biochemist
                                      Dr. Silke Wittemann, biochemist




                                                                             NMI Annual Report 2002/03   49
                                                            Publications




Publications
Bauerdick S., Burkhardt C., Kern D.P., Nisch W.             Stett, A., Bucher, V., Burkhardt, C., Weber, U., Nisch,      Sachs, H.G., Framme, C., Gekeler, F., Schanze, T.,
(2003). Substrate-integrated microelectrodes with           W. (2003). Patch-clamping of primary cardiac cells           Nisch, W., Brunner, U., Zrenner, E., Gabel, V.P. (2003).
improved charge transfer capacity by 3-dimensional          with micro-openings in polyimide films. Med Biol Eng         Surgical Procedure for the Implantation of Foil
micro-fabrication. Biomedical Microdevices 5 (2),           Comput 41, 233-240.                                          Strip Stimulators for Repentable Stimulation in the
93-99.                                                                                                                   Subretinal Space of Cats for the Purpose of Stimula-
                                                            Stett, A., Burkhardt, C., Weber, U., van Stiphout, P.,       tion Parameter Determination in Visual Prosthesis
Bauerdick S., Burkhardt C., Rudorf R., Barth W.,            Knott, T. (2003). CYTOCENTERING: a novel technique           Development. Invest. Ophthalmol. Vis. Sci. 44,
Bucher V., Nisch W. (2003). In-situ monitoring of           enabling automated cell-by-cell patch clamping with          E-Abstract 5073.
electron beam induced deposition by atomic force            the CYTOPATCH chip. Receptors Channels 9, 59-66.
microscopy in a scanning electron microscope. Micro-                                                                     Schwenk, J.M., Stoll, D., Templin, M.F., Joos, T.O.
electronic Engineering 67-68, 963-969.                      Stett, A., Egert, U., Guenther, E., Hofmann, F., Meyer,      (2002). Cell microarrays: an emerging technology
                                                            T., Nisch, W., Hämmerle, H. (2003). Biological appli-        for the characterization of antibodies. Biotechniques
Breisch, S., de Heij, B., Löhr, M., Stelzle, M. (2004).     cation of microelectrode arrays in drug discovery and        Suppl, 54-61.
Selective chemical surface modification of fluidic          basic research. Anal Bioanal Chem 377, 486-495.
microsystems and characterization studies. J. Micro-                                                                     Shinoda, K., Kobuch, K., Gekeler, F., Nisch, W., Sailer,
mechanics and Microengineering 14, 497-505.                 Stoll, D., Templin, M.F., Schrenk, M., Traub, P.C.,          H., Blatsios, G., Eckert, E., Gabel, V.P., Zrenner, E.
                                                            Vöhringer, C.F., Joos, T.O. (2002). Protein microarray       (2003). Long Term Retinal Function after Repeated
Brockmann, W., Geiss P.L., Klingen J., Schröder B.          technology. Front Biosci 7, c13-32.                          Ab Externo Implantation and Explantation of Subreti-
(2004). Klebtechnik. In: P.e.D.W.K. Dr. Ing. Roland                                                                      nal Electronic Devices in Rabbit. Invest. Ophthalmol.
Dittmeyer (Ed.) Chemische Technik - Prozesse und            Templin, M.F., Stoll, D., Schrenk, M., Traub, P.C.,          Vis. Sci. 44, E-Abstract 5076.
Produkte, Industrieprodukte, Kapitel 7, Band 7              Vöhringer, C.F., Joos, T.O. (2002). Protein microarray
(WILEY-VCH. Weinheim).                                      technology. Trends Biotechnol 20, 160-166.                   Stett, A., Herrmann, T., Hofmann, F., Guenther,
                                                                                                                         E. (2003). Electrophysiological Cell And Tissue
Bucher, V., Brugger, J., Kern, D., Kim, G.M., Schubert,     Templin, M.F., Stoll, D., Schwenk, J.M., Potz, O.,           Based Assays For Drug Testing With Microelec-
M., Nisch, W. (2002). Electrical properties of light-       Kramer, S., Joos, T.O. (2003). Protein microarrays:          trode Arrays. http://www.sbsonline.org/03talks/
addressed sub-µm electrodes fabricated by use of            promising tools for proteomic research. Proteomics           030401213456.htm
nanostencil-technology. Microelectronic Engineering         3, 2155-2166.
61-62, 971-980.                                                                                                          Stett, A., Knott, T., Polder, H.R., Nisch, W. (2002).
                                                            van Bergen, A., Papanikolaou, T., Schuker, A., Möller,       Automated Patch-Clamp Approach Providing High-
Dürr, M., Kentsch, J., Muller, T., Schnelle, T., Stelzle,   A., Schlosshauer, B. (2003). Long-term stimulation of        Content Screening. Biophys.J. (Annual Meeting
M. (2003). Microdevices for manipulation and accu-          mouse hippocampal slice culture on microelectrode            Abstract), 266a.
mulation of micro- and nanoparticles by dielectro-          array. Brain Res Brain Res Protoc 11, 123-133.
phoresis. Electrophoresis 24, 722-731.                                                                                   Zrenner, E., Gekeler, F., Stett, A., Nisch, W., Sachs,
                                                            Abstracts                                                    H.G., Gabel, V.P., Kisvarday, Z.F., Eysel, U., Schanze,
Hämmerle, H., Kobuch, K., Kohler, K., Nisch, W.,            Angres B., Kuschel C., Steuer H., Stumpf S., Benz K.,        T., Eckhorn, R., MPDA-Consortium (2003). Subretinal
Sachs, H., Stelzle, M. (2002). Biostability of micro-       Stoop R., Hämmerle H. (2003). Multiple Substrate             Microphotodiode Arrays: A Promising Road to Resti-
photodiode arrays for subretinal implantation. Bioma-       Array (MSA): A screening tool for regulation of              tution of Vision in Degeneration of the Outer Retina.
terials 23, 797-804.                                        cell function and adhesion by extracellular matrix.          Invest. Ophthalmol. Vis. Sci. 44, E-Abstract 4204.
                                                            www.sbsonline.org/03poster/030527130911.htm
Helle, T., Deiss, S., Schwarz, U., Schlosshauer, B.                                                                      Non reviewed articles
(2003). Glial and neuronal regluation of the lipid car-     Blatsios, G., Bondzio, L., Sailer, H., Shinoda, K.,          Guenther, E., Stett, A., Meyer, T. (2003). Mikroele-
rier R-F ABP. Experimental Cell Research, 88-97.            Kobuch, K., Sachs, H.G., Gekeler, F., Nisch, W.,             ktroden-Array basierte Biosensoren. Biospektrum 6,
                                                            Gabel, V.P., Zrenner, E. (2003). Temperature Change          772-774.
Joos, T.O., Stoll, D., Templin, M.F. (2002). Miniatur-      in the Subretinal Space During Infrared Irradiation.
ised multiplexed immunoassays. Curr Opin Chem Biol          Invest. Ophthalmol. Vis. Sci. 44, E-Abstract 5077.           Joos, T.O., Bodovitz, S. (2003). Multidimensional
6, 76-80.                                                                                                                proteomics. Drug Discov Today 8, 879-880.
                                                            Gabel, V.P., Sachs, H.G., Gekeler, F., Schanze, T.,
Kentsch J., Dürr M., Schnelle T., Gradl G., Müller T.,      Hesse, L., Eysel, U., Kisvarday, Z.F., Nisch, W., Brunner,   Knott T., Single S., Stett A. (2002). Automatisiertes
Jäger M., Normann A., M., S. (2003). Microdevices           U., Zrenner, E. (2002). Subretinal Implant Surgery           Patch-Clamping - Lösungen und Herausforderungen.
for separation, accumulation, and analysis of biologi-      in a Series of 26 Cat Eyes to Prove Evidence of              Laborwelt IV/2002, 20-22.
cal micro- und nanoparticles. IEE Proc.-Nanobiotech-        Cortical Activation. Invest. Ophthalmol. Vis. Sci. 43,
nologie 150, 82-89.                                         E-Abstract 2845.                                             Roth, K., Heeg, P., Reichl, R. (2002). Hygienische
                                                                                                                         Aspekte bei der Aufbereitung von Einwegproduk-
Roth, K., Heeg, P., Reichl, R. (2002). Specific Hygiene     Gora, F.P., Sachs, H.G., Schanze, T., Gekeler, F.,           ten für die Laparoskopische Chirurgie. Krh.-Hyg. +
Issues Relating to Reprocessing and Reuse of Single-        Brunner, U., Gmeiner, H., Stelzle, M., Haemmerle,            Inf.verh. 24 1, 3-8.
Use Devices for Laparoscopie Surgery. Surg Endosc           H., Zrenner, E., Gabel, V.P. (2002). Surgical Subretinal
16, 1091-1097.                                              Implant Exchange In The Cat Model; First Results.            Schwenk J., Pötz O. (2003). Cell Microarrays Present
                                                            Invest. Ophthalmol. Vis. Sci. 43, E-Abstract 4465.           and Future Applications. Screening 4, 36-37.
Schlosshauer, B., Müller, E., Schröder, B., Planck, H.,
Müller, H.W. (2003). Rat Schwann cells in bioresorb-        Gründer T., Fischer H., Benz K., Gonser B., Fink U.,         Stelzle M., Kentsch J., Schnelle T., Müller T., Gradl G.,
able nerve guides to promote and accelerate axonal          Gaissmaier C., Hämmerle H., Stoop R. (2003). Col-            Haage A., Normann A., Geggier P., Jäger M. (2003).
regeneration. Brain Res 963, 321-326.                       lagen scaffolds for the treatment of degenerative            Dielektrophoretische Trennung biologischer Mikro-
                                                            cartilage lesions. Osteoarthr. Cart. 11 Suppl. A., 67.       und Nanopartikel. Laborwelt 4, 12-13.
Schlosshauer, B., Steuer, H. (2002). Comparative
Anatomy, Physiology and In Vitro Models of the              Kobuch, K., Stelzle, M., Kohler, K., Shinoda, K.,            Stoll, D., Bachmann, J., Templin, M.F., Joos, T.O.
Blood-Brain and Blood-Retina Barrier. Curr. Med.            Gekeler, F., Hämmerle, H., Zrenner, E., Gabel, V.P.          (2004). Microarray technology: an increasing variety
Chem. - Central Nervous System Agents 2, 175                (2003). Biostability and Biocompatibility of Subretinal      of screening tools for proteomic research. TARGETS
- 186.                                                      Electronic Devices after Subretinal and Subcutaneous         3, 24-32.
                                                            Implantation in Rabbits. Invest. Ophthalmol. Vis. Sci.
Steinbach, K., Volkmer, H., Schlosshauer, B. (2002).        44, E-Abstract 5075.                                         Templin, M.F., Stoll, D., Schrenk, M., Traub, P.C.,
Semaphorin 3E/collapsin-5 inhibits growing retinal                                                                       Vöhringer, C.F., Joos, T.O. (2002). Protein microarray
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                                                            M., Hämmerle H., Stoop R. (2003). Peptide-based
                                                            Multi-Substrate Array (MSA): a novel tool to investi-
                                                            gate chondrocyte regulation by extracellular matrix.
                                                            Osteoarthr. Cart 11 Suppl. A., 96.
50   NMI Annual Report 2002/03
Projects




Projects supported by the

BMBF (Federal Ministry of Education and Research)

16SV1744                 Multiwell Microsensor Plate for cell-based drug testing - MiMP
16SV1769                 Microfluidic processor for analysis of cellular functions
16SV1644                 Development of high performance microdrives with new powderbased
                         manufacturing processes
0311962                  Tissue engineering of neural implants for pain treatment
0312879A                 pepART - Peptide based Proteome-Analysis by Array Technology :
                         development and validation of a novel parallelised analytical method
01GN0111                 Microstructured bioresorbable nerve guide with glial cells for nerve
                         regeneration
0311568                  Growth factors and celladhesion molecules for regeneration of peripheral
                         neurons
13N7939                  Nanopatternd electrode system for manipulation of biological micro- and
                         nanoparticels
0310964A                 Use of the microelectrode array technology in the pharmaceutical
                         biotechnology
13N8004                  Site specific preparation of samples for transmission electron microscopy
                         (TEM)
01KP0009                 Retina implant
0311532/9                Microelectrode implant
0311538                  Pigmented retina epithelium as blood-brain barrier model
0312995                  Bioprofile: Therapeutic use of adult mesenchymal stem cell for cartilage-
                         and bone tissue engineering
0312626                  New procedure for the production of autologous tissue replacement for
                         orthopedic applications


Baden-Württemberg

4-4332.62-NMI/17         Technologies for Medical Products with Improved Functional Properties
4-4332.65-NMI/15         Chip-based genome-, proteome-, and cellome analysis for the
                         regenerative medicine
431-1-7/2                Bio-disc: CD-based diagnostic system
4-4331.1/67              Research and prevention of damages on lubricated machinery elements
                         under practical conditions


european union

G6ST-CT-2001-50121       Cleantest CRAFT - 1999-70573 Testing and Measurement Procedure for
                         the Validation of the Cleaning Behaviour of Reusable Surgical Devices
QLK6-CT2000-00487        Spare Parts - Chondral and Osseous Tissue Engineering www. tissue-
                         engineering-oc.com


other projects

DBU 15418                Development of definied soluble and industrial usable adhesives for
                         environmental recycling of adhesive bondings and for improvement of
                         ecological balances
FVA 289                  Characterization of tribolayers II


The NMI thanks the Federal Ministry of Education and Research (BMBF), the Ministry of Economic
Affairs of Baden-Württemberg, the Landesstiftung foundation Baden-Württemberg and the
European Union for support of the obove projects.




                                                                                             NMI Annual Report 2002/03   51
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52   NMI Annual Report 2002/03
NMI Natural and Medical Sciences Institute
at the University of Tübingen
Markwiesenstraße 55
D-72770 Reutlingen
Phone: +49 7121 515 30-0
                                             www.mees-zacke.de




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