NanoDLSay_ A New Platform Technology for Biomolecular Detection .._1_ by malj


									NanoDLSayTM – A New Solution for
Biomolecular Detection and Analysis

         February 2010 Copyright of Nano Discovery, Inc.
Biomolecular Detection and Analysis:
the foundation of the whole modern life
science research and medical diagnosis

          February 2010 Copyright of Nano Discovery, Inc.
What is NanoDLSayTM: Gold Nanoparticle Coupled with
  Dynamic Light Scattering (DLS) for Biomolecular Assay

           Y                 Y                                            Y                Y






           Y                 Y           antigen                          Y                Y

           Gold nanoparticle-
          antibody conjugates
                                                                              Size increase

                         Assay Procedure
                                                                              Increasing antigen concentration

      Add                Incubate            Measure
     sample                                  by DLS

                                                                                     50         100 Size (nm)
                     February 2010 Copyright of Nano Discovery, Inc.
       Why Gold Nanoparticles (GNPs)?
     Gold nanoparticles (GNPs) have exceptionally large light scattering cross
      section at or near their surface plasmon wavelength region
     Gold nanoparticles scatter light 105 times stronger than a fluorescent dye
      molecule; 100s times stronger than polystyrene (PS) latex particles
     The scattered light of gold nanoparticles does not suffer from the
      photobleaching often encountered in fluorescent molecules
     Detection limit of DLS for GNPs can easily reach fM to aM range

A                                 B                                   C

                                       PS particle
                                                                        Gold nanorods

    Figure: Dark field optical images of GNPs mixed with human serum (A) and PS particles (B)

                           February 2010 Copyright of Nano Discovery, Inc.
                          Dynamic Light Scattering (DLS)
                                                       Detector     Scattered light intensity fluctuation
Laser beam

                             Sample solution

                          Particle size and size distribution
 Intensity Distribution

                                                                    Autocorrelate intensity fluctuations

                               50       100     150
                            Average particle size (nm)
Unique Features of NanoDLSayTM
       Obtain results in minutes instead of hours and days
       Easy to conduct (a one-step process!)
       Almost labor-free, no special training needed
       Simple instrument ($40-60K, instead of $100sK +)
       Low cost and high sensitivity
       Can be easily adapted for protein panel analysis
       Extensive range of applications

Comparison with Traditional Techniques:
   ELISA: takes days to prepare and hours to conduct the assay
   Western blot: takes days to complete, labor-intensive, special training
   Surface plasmon resonance: too expensive ($200-500K)
   Applications: limited
                                          February 2010 Copyright of Nano Discovery, Inc.
Various Types of Gold Nanoparticle Size Change Upon
        Binding with Target Protein Molecules
                                                      Sandwich assay

                                                      interaction study

                                                    Protein complex/
                                                   aggregate detection

                                                    Particle size
 February 2010 Copyright of Nano Discovery, Inc.
Broad Applications of NanoDLSayTM
     General assay for protein detection & analysis
     Protein-protein interaction study
     Biomolecular binding kinetics study
     Receptor-ligand identification
     Antibody isotyping and quality control analysis
     Protein complex analysis
     Protein aggregation study
     Detection of non-protein biomolecules
     Detection of small chemicals and ions
     Protein inhibitor screening and drug development
     Biopharmaceutical research and development
     Detection of viruses and bacteria
     Nanoparticle bioconjugate development
     Nanoparticle quality control
                                          February 2010
     Nanoparticle size analysis          Copyright of Nano Discovery, Inc.
                              Application 1. As a General Sandwich
                              Immunoassay for Protein Detection
                                  Y                 Y                               Y             Y






                                  Y                 Y        Target protein         Y             Y

                                            Huo, et al. JACS, 2008, 130, 2780-2782

                                                                    •   Two different monoclonal antibodies are
Average particle size (nm)

                                                                        conjugated to two different GNPs
                                                                    •   Sandwich structure formation between the
                                                                        target protein and two GNP probes will
                                                                        cause nanoparticle cluster formation,
                                                                        therefore, lead to the average particle size
                                                                        increase of the assay solution
                                                                    •   The average particle size increase can be
                                                                        correlated to target protein concentration
                                                                    •   Two monoclonal antibodies may be
                                                                        replaced by a polyclonal antibody
                             Target protein concentration

                                           February 2010 Copyright of Nano Discovery, Inc.
                               Application 2. Monitor Gold Nanoparticle
                              Bioconjugation Process and Quality Control
                                                                     •   An extremely powerful tool to monitor gold
                                                                         nanoparticle bioconjugation process
                                                                     •   Analyze the quality, stability and binding affinity
                                                                         of gold nanoparticle bioconjugates
                             Nanoparticle size increase
                                                                     Huo, et al. Anal. Chem. 2009, 81, 9425-9432
Average particle size (nm)

                                                                                Average particle size (nm)
                                          In-situ monitoring of the                                               Antibody concentration
                                             adsorption process                                                        effect study

                                         Incubation Time (min)                                               Antibody Concentration (µg/mL)

                                                     February 2010 Copyright of Nano Discovery, Inc.
                                         Application 3. Protein-Protein
                                         Binding and Interaction Study
                                                                            •   In-situ monitoring of protein-protein
                             Y                              Y                   binding and interaction study
                                                                            •   When target protein binds to protein
                                                                                conjugated to gold nanoparticles, the


                             Y         Matching             Y                   particle size will increase
                                     target protein                         •   A function very similar to the Surface
                                                                                Plasmon Resonance technique
                                                                            •   Example 1: can be used to confirm the
Average particle size (nm)

                                                                                binding affinity of bioconjugated gold
                                                                            •   Example 2: can be used for antibody
                                                                                isotyping and quality control analysis

                                                                                                        No interaction

                                                                                Y                       No size increase
                                      Incubation Time (min)

                                              Huo, et al. American Biotechnology Laboratory 2010, in press
                                                      February 2010 Copyright of Nano Discovery, Inc.
                                             Application 4. Detect Protein
                                            Complex/Aggregate Formation
How to identify protein complex/aggregate formation from NanoDLSay analysis:
                             •   Nonlinear increase of nanoparticle size at a critical target protein concentration
                             •   The particle size increases dramatically and quickly at this particle concentration
                             •   Particle size distribution curve often reveals very broad and multi-model polydispersed
                                 distribution. Run-to-run variation is often large
                                                                                               Size distribution curve
                                       Dose-Response Curve
Average particle size (nm)

                                                                                   Relative Intensity

                                  Target protein concentration                                          Size distribution (nm)

                                                    February 2010 Copyright of Nano Discovery, Inc.
      Examples: Refer to Publications
1)   Jans, H.; Liu, X.; Austin, L.; Maes, G.; Huo, Q. Dynamic light scattering as a
     powerful tool for gold nanoparticle bioconjugation and biomolecular binding
     study. Anal. Chem. 2009, 81, 9425-9432
2)   Austin, L.; Liu, X.; Huo, Q. An immunoassay for monoclonal antibody
     isotyping and quality analysis using gold nanoparticles and dynamic light
     scattering. American Biotechnology Laboratory 2010, in press
3)   Liu, X.; Huo, Q. A washing-free and amplification-free one-step
     homogeneous assay for protein detection using gold nanoparticle probes and
     dynamic light scattering. J. Immunol. Method 2009, 349, 38-44.
4)   Dai, Q.; Liu, X.; Coutts, J.; Austin, L.; Huo, Q. A one-step highly sensitive
     method for DNA detection using dynamic light scattering. J. Am. Chem. Soc.
     2008, 130, 8138-8139.
5)   Liu, X.; Dai, Q.; Austin, L.; Coutts, J.; Knowles, G.; Zou, J.; Chen, H.; Huo, Q.
     A One-step homogeneous immunoassay for cancer biomarker detection using
     gold nanoparticle probes coupled with dynamic light scattering. J. Am. Chem.
     Soc. 2008, 130, 2780-2782. (also featured at JACS Select #5, 2009, free)
6)   Ray, P.C. et al. Use of gold nanoparticles in a simple colorimetric and
     ultrasensitive dynamic light scattering assay: selective detection of arsenic in
     groundwater. Angew. Chem. Int. Ed. 2009, 48, 9668-9671.
For Further Information


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Tel: 407-770-8954

12565 Research Parkway Suite 300, Orlando, FL 32826

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