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Purification and Analysis of Strep-tagged Antibody-Fragments

  • Martin Schlapschy
  • Arne SkerraEmail author
Protocol
Part of the Springer Protocols Handbooks book series (SPH)

Abstract

The Strep-tag is a nine-residue minimal peptide sequence with intrinsic affinity towards streptavidin. This peptide – or its improved version Strep-tag II – can be fused to recombinant antibody fragments and other proteins in various fashions. The fact that the interaction between the Strep-tag and streptavidin is highly specific and can be reversed upon addition of biotin (or suitable derivatives such as desthiobiotin) enables a highly efficient affinity chromatography under biochemically mild conditions. Here we describe a protocol for the production and purification of Strep-tagged antibody fragments from bacterial cell lysates using affinity chromatography on a matrix carrying an engineered streptavidin (Strep-Tactin), followed by analysis in ELISA and on Western blots using Strep-Tactin enzyme conjugates or anti-Strep-tag antibodies. Thus, the Strep-tag, which is short, biologically inert, proteolytically stable and does not interfere with membrane translocation or protein folding, offers a versatile tool both for the rapid isolation of antibody fragments and for their detection or molecular analysis.

Keywords

Antibody Fragment Host Cell Protein Periplasmic Extraction Biotin Carboxyl Carrier Protein Recombinant Antibody Fragment 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Springer-Verlag Berlin Heidelberg 2010

Authors and Affiliations

  1. 1.Lehrstuhl für Biologische ChemieTechnische Universität MünchenFreising-WeihenstephanGermany

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