Application of bispecific antibody against antigen and hapten for immunodetection and immunopurification.
- Author:
Hyori KIM
1
;
Sunyoung PARK
;
Hwa Kyoung LEE
;
Junho CHUNG
Author Information
1. Department of Biochemistry and Molecular Biology, Seoul National University College of Medicine, Seoul, Republic of Korea. jjhchung@snu.ac.kr
- Publication Type:Original Article ; Research Support, Non-U.S. Gov't
- Keywords:
bispecific antibody;
hapten;
immunoblot;
immunoprecipitation
- MeSH:
Animals;
Antibodies, Bispecific/*immunology;
HEK293 Cells;
Haptens/*immunology;
Humans;
Immunoblotting/*methods;
Immunoprecipitation/methods;
Rabbits;
Single-Chain Antibodies/immunology
- From:Experimental & Molecular Medicine
2013;45(9):e43-
- CountryRepublic of Korea
- Language:English
-
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.
