摘要
目的探讨重组质粒pET22b-SEA经原核表达后,金黄色葡萄球菌肠毒素A(SEA)对人外周血单核细胞(PBMC)杀瘤活性的影响。方法将重组质粒pET22b-SEA转化至E.coli BL21(DE3)扩大培养,IPTG诱导后,进行SDS-PAGE分析;密度梯度离心分离PBMC,不同浓度SEA诱导PBMC,12、36、60h后用MTT法测定PBMC对人肺腺癌细胞A549的杀伤活性。结果经诱导,pET22b-SEA得到表达,产物相对分子质量约为30kDa,与理论值大小一致;随着SEA浓度的升高PBMC的杀瘤活性显著增强,在36h、高效靶比、高SEA浓度的情况下,杀伤率最高可达57.21%。结论重组质粒pET22b-SEA在大肠杆菌中成功表达;SEA可诱导PBMC对人肺腺癌细胞A549产生更强的杀瘤活性。
Objective To investigate the effect of staphylococcus enterotoxin A （SEA）on the peripheral blood mononuclear cell（ PBMC）to kill lung adenocarcinoma, after recombinant plasmid pET22b-SEA is expressed in E. coll. Methods Recombinant plasmid pET22b-SEA was transformed into E. coli BL21 （ DE3 ） and cultured, SDS-PAGE analysis was performed after IPTG induction, and the PBMC was separated by density gradient centrifugation,and PBMC was induced by the different concentrations of SEA, and the killing effect of PRMC on human lung adenocarcinoma cells-AS49 was detected by MTT after 12h,36h,60h. Results After IPTG induced, pET22b-SEA was expressed, and the relative molecular mass of the production was about 30KDa which was in accordance with the theoretical value ; the killing tumor effect of PBMC was remarkably enhanced with the increase of SEA concentration. The killing rate was 57.21% under the circumstance of high high ratio of effect vs target, high SEA concentration, after 36h. Conclusion Recombinant plasmid pET22b-SEA is expressed successfully in E. coli, and SEA can induce PBMC to produce stronger killing tumor effect on human lung adenocarcinoma cells-A549.
出处
《河北医药》
CAS
2010年第1期3-5,共3页
Hebei Medical Journal
基金
天津市科委资助应用基础研究重点项目(编号:06YFJZJC02600)
