Novel Protein Drug Delivery System (PowerPoint download)

							  Small Diameter Centrifuge
            Tube
                   Team Members:
            Sarajane Stevens - Team Leader
              Anita Zarebi - Communicator
             Christopher Westphal - BWIG
                  Katy Reed - BSAC


     Client:                          Advisor:
Pamela Kling, M.D.            Professor Kristyn Masters
Presentation Outline
 Problem Statement
 Background Information
 Research Competition
 Suggested Materials
 Design Constraints
 Alternative Designs
 Future Work
          Problem Statement
Client: Pamela J. Kling, M.D.
Blood samples <1ml
Small Diameter Centrifuge Tube
                                 plasma
 Super clear, strong plastic

 Large length:diameter ratio

 Volume ~ 70μl                  WBC
 Adaptor for centrifuge
                                 RBC
      Background Information
Anemia in newborns
   Red blood cell
    deficiency
   Drug research
   Multiple, consistent
    samples
Reticulocytes
   Immature red blood
    cells
                           R=Reticulocyte (rounded)
   Stimulates cell
                           E=Red Blood Cell
    production             (biconcave)
   Analyzed for iron
      Background Information
                                 Cord Blood Sample
                                  collected at birth


                               Brought to Blood Bank
                               (stored at 4C, 4 Center)


                              To be discarded at 7 days


                        Picked up from 4 Center by Kling Lab


                               Centrifuged at 3000 rpm
                                  Plasma removed


                                 Washed in PBS x3
                               Make Hct 50% with PBS


             No Enrichment                                Retic Enrichment Step
                                                               BME Method


ZnPP/H measured x2    Retiulocyte Smear      ZnPP/H measured x2        Retiulocyte Smear
  on 20 µL aliquots   stained with BCB        on 20 µL aliquots         stained with BCB


                        Count Retics                                     Count Retics
                       Microscopically                                  Microscopically
Current Devices: Centrifuge Tubes

  Typical blood sample
  ~10ml
  Smaller volumes
     Small length:diameter
      ratio
  Material deforms in
  floor model centrifuge
Research Competition
           Private research
           companies
               Techniques, products
                for centrifuge systems
           Direct use
               Beckman-Coulter
               Fisher Scientific
               Eppendorf
Suggested Materials
           Polyethylene
              Deforms in floor model
              Cloudy
           Polypropylene
              Strong
              Clear
           Tygon tubing
              Cut to desired length
           Superclear plastics
           Polycarbonate
Design Constraints

Super clear plastic
Volume <1ml
Length:Diameter ratio ~ 50:1
1 hour long centrifugation
15,000-37,000 x g
Temperature 30°-40°C
           Problem Overview
      Separation           Extraction

Trial & Error Options   Seal and Cut
   Microhematocrit
    centrifuge          Removal by pipet

   Ultracentrifuge     “Titration” method
   Materials

   Tube choice
Design I: Seal and Cut




 Critoseal
 Hematocrit tube cutter
 Collection container or syringe
      Design I: Pros and Cons
Pros                             Cons
    Quick and easy                 Use more Critoseal
    Increases purity of            Remnants of tubes
     sample                          create untidiness
    Utilizes current lab
     equipment


Critoseal          Whole Blood   Reticulocytes   Critoseal
Design II: Simple Extraction
       Syringes/Pipette

        Narrow tubes
        Tube holder
        Magnifying glass
 Design II: Pros and Cons
               Syringe/Pipette
Pros                             Cons
   Syringe/Pipette                 Syringe
       Variable volume                 Inconvenient
       No vacuum                       withdrawal
   Syringe                         Pipette
       Multiple in one syringe         One tube at a time
   Pipette
       One hand operation
Design III: Titration
    Design III: Pros and Cons
Pros                       Cons
   Eliminates suction        Direct contact with
   Ability to stop flow       sample--gloved hand
    early                     Multiple tests--time
   Isolate/pipet easily       consuming
                              Tubes may break
                               upon slicing
        Future Work
Test all methods--combine?
Plastics research
   Temperature tolerance
   High “g” tolerance
Construct AND test prototype
Consult client
                                References
[1] General Use Needles. 15 Feb 2005.
     http://catalog.bd.com/bdCat/listProducts.doCustomer?categoryID=134&priority=25&categoryN
     ame=General%20Use%20Needles&categoryImgPath=/ecat/images/f07/family/needles.jpg&sor
     tByField=Product%20Number&viewPageNum=0
[2] GELoader® Tips. 13 Feb 2005. http://www.eppendorf.com/script/cat-
     prd.php?l=25&region=&b=9734
[3] Diabetes Dictionary. 13 Feb 2005. http://diabetes.niddk.nih.gov/dm/pubs/dictionary/P-T.htm
[4] Murphy, J.R. 1973. Influence of temperature and method of centrifugation on the separation of
     erythrocytes. Journal of Laboratory Clinical Medicine 82;334-341
[5] Brugnara, C. et al. 1993. Red blood cell regeneration induced by subcutaneous recombinant
     erythropoietin: iron-deficient erythropoiesis in iron-replete subjects. Blood. 1993; 81(4): 956-964
[6] SuperClear™ Microcentrifuge Tubes. 17 Feb 2005.
     http://www.labdepotinc.com/product_details~id~52~pid~150.aspx
[7] VisiPak Extrusions. 17 Feb 2005. http://www.visipak-extrusion.com/
[8] Fisherbrand Plastic Microhematocrit Tubes. 17 Feb 2005.
     https://www1.fishersci.com/Coupon?gid=178612&cid=1333
[9] Hematology and Immunology Studies. 17 Feb 2005. http://history.nasa.gov/SP-368/p212.jpg
[10] Axeda. 11 Feb 2005. http://www.axeda.com/images/customers/BeckmanCoulter_logo.jpg
[11] Chem Lab Cricket Team. 13 Feb 2005. http://www-leeper.ch.cam.ac.uk/Fisher2.jpg
[12] BioBank. 12 Feb 2005. http://www.biobank.co.kr/maker/eee/eppendorf-logo.gif
Questions?