Preparation and Characterization of a Polyclonal Antibody against Human Actin Filament-Associated Protein-120 kD
AbstractActin filament-associated protein-120kD (AFAP-120) is an alternatively spliced isoform of actin filament-associated protein-110kD (AFAP-110) and contains an additional neuronal insert (NINS) fragment in addition to identical domains to the AFAP-110. Unlike AFAP-110 widely expressed in tissues, AFAP-120 is specifically expressed in the nervous system and plays a role in organizing dynamic actin structures during neuronal differentiation. However, anti-AFAP-120 antibody is still commercially unavailable, and this may hinder the function research for AFAP-120. In this study, we simultaneously used the ABCpred online server and the BepiPred 1.0 server to predict B-cell epitopes in the exclusive NINS sequence of human AFAP-120 protein, and found that a 16aa-peptide sequence was the consensus epitope predicted by both tools. This peptide was chemically synthesized and used as an immunogen to develop polyclonal antibody against AFAP-120 (anti-AFAP-120). The sensitivity and specificity of anti-AFAP-120 were analyzed with immunoblotting, immunoprecipitation, and immunofluorescence assays. Our results indicated that anti-AFAP-120 could react with over-expressed and endogenous human AFAP-120 protein under denatured condition, but not with human AFAP-110 protein. Moreover, native human AFAP-120 protein could also be recognized by the anti-AFAP-120 antibody. These results suggested that the prepared anit-AFAP-120 antibody would be a useful tool for studying the biochemical and biological functions of AFAP-120. View Full-Text
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Chen, Y.; Liu, Y.; Guo, J.; Tang, T.; Gao, J.; Huang, T.; Wang, B.; Liu, S. Preparation and Characterization of a Polyclonal Antibody against Human Actin Filament-Associated Protein-120 kD. Int. J. Mol. Sci. 2016, 17, 942.
Chen Y, Liu Y, Guo J, Tang T, Gao J, Huang T, Wang B, Liu S. Preparation and Characterization of a Polyclonal Antibody against Human Actin Filament-Associated Protein-120 kD. International Journal of Molecular Sciences. 2016; 17(6):942.Chicago/Turabian Style
Chen, Yujian; Liu, Yong; Guo, Jiayu; Tang, Tao; Gao, Jian; Huang, Tao; Wang, Bin; Liu, Shaojun. 2016. "Preparation and Characterization of a Polyclonal Antibody against Human Actin Filament-Associated Protein-120 kD." Int. J. Mol. Sci. 17, no. 6: 942.
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