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Identification of Target and Function Specific Antibodies for Effective Drug Delivery

  • Yu Zhou
  • James D. Marks
Protocol
Part of the Methods in Molecular Biology™ book series (MIMB, volume 525)

Abstract

Phage antibody technology is a powerful approach for generating human antibodies to virtually any target antigen. For many therapeutic applications, it is useful to generate antibodies that bind to cell-surface receptors in a manner where binding results in internalization of the antibody. This allows use of the antibody to deliver toxic payloads intracellularly to achieve a therapeutic effect. Here we describe how phage antibody libraries can be directly selected on tumor cell lines to generate antibodies binding cell-surface receptors and which are rapidly internalized upon binding. Protocols are provided showing how to (1) directly select internalizing antibodies from phage antibody libraries; (2) screen phage antibodies in a high-throughput flow cytometry assay for binding to the tumor cell line used for selection; (3) identify the antigen bound by the phage antibody using immunoprecipitation and mass spectrometry; and (4) verify and quantitate such that phage antibodies are internalized.

Key words

Phage antibody antibody internalization targeted drug delivery cell selection flow cytometry 

Notes

Acknowledgments

This work was partially supported by NIH grant P50 CA58207.

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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Yu Zhou
    • 1
  • James D. Marks
    • 1
  1. 1.University of CaliforniaSan FranciscoUSA

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