Construction,expression and in vitro specific killing activity of a recombinant fusion protein:anti-CD20Fab-LDM
- VernacularTitle:重组融合蛋白抗CD20Fab-LDM的构建、表达及体外活性研究
- Author:
Xin CHENG
;
Ming YANG
;
Dongmei FAN
;
Yuanfu XU
;
Yuan ZHOU
;
Yingdai GAO
;
Jinhong WANG
;
Yuan ZHOU
;
Wei LI
;
Dongsheng XIONG
- Publication Type:Journal Article
- Keywords:
CD20;
LDM;
gene engineered antibody;
immunotherapy;
non-Hodgkin′s lymphoma Biotoxins(NHL);
biotoxins
- From:
Chinese Pharmacological Bulletin
2009;25(12):1650-1654
- CountryChina
- Language:Chinese
-
Abstract:
Aim To construct and express anti-CD20Fab-LDP,generate anti-CD20Fab-LDM and identify its biological activity.Methods PCR and overlapping PCR were used to construct anti-CD20Fab-LDP.DNA sequence was analyzed by the Terminus of Dideoxy Nucleotide.The product was purified by affinity chromatography and analyzed by Western blot and its antigen-binding activity was examined by FACS.Specific killing activity in vitro of anti-CD20Fab-LDM was analyzed by MTT.Results The data of DNA sequence showed that anti-CD20Fab-LDP was correct.The fusion protein was recovered in high yield(up to 4 mg·L~(-1))after proteinG purification.The fusion protein could bind to Raji cells(CD20+),and similar affinity data were obtained with anti-CD20Fab.Anti-CD20Fab-LDP showed potent cytotoxicity to Raji cells with IC_(50) values of 0.9×10~(-10) mol·L~(-1).Conclusions Anti-CD20Fab-LDP with high level expression was successfully obtained and could bind to Raji cells cells.Anti-CD20Fab-LDM showed specific killing activity to Raji cells in vitro.
