Exp Mol Med.
2013 Sep;45(9):e43.
Application of bispecific antibody against antigen and hapten for immunodetection and immunopurification
- Affiliations
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- 1Department of Biochemistry and Molecular Biology, Seoul National University College of Medicine, Seoul, Republic of Korea. jjhchung@snu.ac.kr
- 2Cancer 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 x 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.
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