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Exp Mol Med. 2013 Sep 27;45:e43. doi: 10.1038/emm.2013.83.

Application of bispecific antibody against antigen and hapten for immunodetection and immunopurification.

Author information

1
1] Department of Biochemistry and Molecular Biology, Seoul National University College of Medicine, Seoul, Republic of Korea [2] Cancer Research Institute, Seoul National University, Seoul, Republic of Korea.

Abstract

We present a bispecific antibody that recognizes an antigen and a hapten and can be applied to various biological assays, including immunoblotting and immunoprecipitation. In immunoblot analysis of serum, an anti-C5 × anti-cotinine bispecific tandem single-chain variable fragment (scFv)-Fc fusion protein and cotinine-conjugated horseradish peroxidase (HRP) generated a clean signal without the high background that was observed in a parallel experiment using HRP-conjugated goat anti-rabbit immunoglobulin G (Fc-specific) antibody. In immunoprecipitation analysis of serum, use of the bispecific tandem scFv-Fc fusion protein and cotinine-crosslinked magnetic beads significantly reduced the amount of protein contaminants compared with a parallel experiment done with protein A agarose beads. In subsequent immunoblot analysis, use of cotinine-HRP as the secondary probe instead of HRP-conjugated goat anti-rabbit IgG (Fc-specific) antibody successfully eliminated the band corresponding to the bispecific tandem scFv-Fc fusion protein.

PMID:
24071736
PMCID:
PMC3789267
DOI:
10.1038/emm.2013.83
[Indexed for MEDLINE]
Free PMC Article

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