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Kinetic Screening in the Antibody Development Process

  • Michael SchrämlEmail author
  • Matthias Biehl
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 901)

Abstract

Kinetic screening is of paramount importance when it is to select custom-made antibodies, tailored for their respective scientific, diagnostic, or pharmaceutical application. Here a kinetic screening protocol is described, using a Biacore A100 surface plasmon resonance biosensor instrument. The assay is based on an Fc-specific antibody capture system. Antibodies from complex mixtures, like from mouse hybridoma supernatants are captured on the sensor surface in an oriented manner. The method uses a single injection of one antigen concentration for the determination of six relevant screening parameters, which comprehensively describe the antibody’s kinetic rate profile and its valence mode. The method enables the scientist to rank and finally select rare and outstanding antibodies according to their kinetic signatures.

Key words

Kinetic screening Molar Ratio Valence Dissociation half-life (t1/2 dissBinding Late Stability Late Antibody Capture Level kd Surface plasmon resonance 

References

  1. 1.
    Canziani GA, Klakamp S, Myszka DG (2004) Kinetic screening of antibodies from crude hybridoma samples using biacore. Anal Biochem 325(2):301–307PubMedCrossRefGoogle Scholar
  2. 2.
    Rich RL, Myszka DG (2007) Higher-throughput, label-free, real-time molecular interaction analysis. Anal Biochem 361(1):1–6PubMedCrossRefGoogle Scholar
  3. 3.
    Wassaf D, Kuang G, Kopacz K et al (2006) High-throughput affinity ranking of antibodies using surface plasmon resonance microarrays. Anal Biochem 351(2):241–253PubMedCrossRefGoogle Scholar
  4. 4.
    Safsten P, Klakamp SL, Drake AW et al (2006) Screening antibody–antigen interactions in parallel using biacore a100. Anal Biochem 353(2):181–190PubMedCrossRefGoogle Scholar
  5. 5.
    Johnsson B, Lofas S, Lindquist G (1991) Immobilization of proteins to a carboxymethyldextran-modified gold surface for biospecific interaction analysis in surface plasmon resonance sensors. Anal Biochem 198(2): 268–277PubMedCrossRefGoogle Scholar
  6. 6.
    Leonard P, Safsten P, Hearty S et al (2007) High throughput ranking of recombinant avian scfv antibody fragments from crude lysates using the biacore a100. J Immunol Methods 323(2): 172–179PubMedCrossRefGoogle Scholar
  7. 7.
    Pasqualucci L, Guglielmino R, Houldsworth J et al (2004) Expression of the aid protein in normal and neoplastic B cells. Blood 104(10): 3318–3325PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Roche Diagnostics GmbHPenzbergGermany

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