Prokaryotic expression of FILIP-1 L and preparation of its polyclonal antibody
10.3969/j.issn.1000-484X.2016.06.014
- VernacularTitle:FILIP-1 L 蛋白的原核表达及多克隆抗体制备
- Author:
Jingfang DU
;
Guoqing DUN
;
Shuman ZHANG
;
Liujie CHU
;
Shulian LI
;
Yanzhong HU
;
Yuanfang MA
- Publication Type:Journal Article
- Keywords:
Filamin A interacting protein 1-like;
Fusion protein;
Polyclonal antibody
- From:
Chinese Journal of Immunology
2016;32(6):832-837
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To express and purify the fusion protein of GST-FILIP-1L and prepare its polyclonal antibody. Methods:The constructed recombinant expression vectors pGEX-4T3-FILIP-1L were transformed into Escherichia Coli BL21. FILIP-1L fusion protein was induced by IPTG and purified by Glutathion Sepharse 4B . The rabbit was immunized by the purified fusion protein,and pro-duced serum with anti-FILIP-1L antibody. The titer of polyclonal antibody was detected by ELISA, the anti-FILIP-1L polyclonal antibody was purified by Active and its combining specificity with FILIP-1L protein was further identified by Western blot. Results:The GST-FILIP-1L fusion protein was highly expressed in E. coli, and its specific polyclonal antibody was obtained after the immunization. The polyclonal antibody purified by Active Ester Agrose was able to combine specially with FILIP-1L protein and transformed FILIP-1L protein in 293 cells and FILIP-1L protein of liver cancer cells, respectively. Conclusion: The GST-FILIP-1L fusion protein was expressed successfully,the anti-GST-FILIP-1L polyclonal antibodies with high titer and specificity are successfully prepared,these antibodies provide an useful experimental tool to research the biological function of the FILIP-1L protein.
