Anti-Histidine Antibodies as Tools for Reversible Capturing of His-Tagged Fusion Proteins for Subsequent Binding Analysis

  • H.-M. Zenn
  • S. Hutschenreiter
  • F. W. HerbergEmail author

The hexahistidine tag is one of most commonly used fusion tags in affinity purification of recombinantly expressed proteins. Real-time binding analysis using Biacore technology allows in-depth characterization of respective association and dissociation patterns of potential binders. Here we tested four commercially available anti-His antibodies for reversible capturing of His-tagged proteins as a basis for a subsequent interaction analysis with non-His-tagged proteins. Anti-penta-, anti-hexa- and anti-RGS-(His)4 antibodies from different distributors were covalently coupled to Biacore sensor chips. Parallel binding studies of 12 heterogeneously sized RGS-(His)6-tagged (Arg-Gly-Ser-(His)6) proteins revealed that the slowest dissociation rate was obtained when using an anti-RGS-(His)4 antibody. Thus in a sandwich binding assay the anti-RGS-(His)4 antibody can be utilized as an appropriate tool for stable yet reversible capturing of RGS-(His)6-tagged proteins with a non-His-tagged protein.


Sensor Chip Dissociation Rate Constant Covalent Coupling Antibody Surface Binding Stoichiometry 
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.



We thank Dr. U. Bierfreund and Dr. W. Jäger from Biacore (GE Healthcare) for support. We thank Dr. H. Seitz (MPI for Molecular Genetics, Berlin, Germany) for providing RGS-(His)6-tagged proteins. F.W.H.'s group is member of Proteome Binders. This work was supported by EU STREP Affinity Proteome to F.W.H.


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Copyright information

© Springer-Verlag 2010

Authors and Affiliations

  1. 1.Department of BiochemistryUniversity of KasselKasselGermany

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