摘要
目的 克隆、表达B7 2 (IgV +C)并进行体外功能测定。方法 用聚合酶链反应 (PCR)技术从B7 2cDNA中克隆B7 2 (IgV +C) ,在此基础上构建表达B7 2 (IgV +C)的原核表达载体pGEX 4T 3 hB7 2 (IgV +C) ;SDS PAGE检测蛋白表达 ,蛋白经变性、复性后在体外协同抗CD3单抗刺激人T细胞 ,3 H TdR掺入法检测T细胞的活化程度。结果 在原核表达载体中成功地克隆了B7 2 (IgV +C) ,SDS PAGE表明在相对分子质量 (Mr) 5 5× 10 3 处有hB7 2 (IgV +C) GST融合蛋白的高效表达 ,其表达量占菌体总蛋白的 33%。体外实验证明 ,此融合蛋白可协同抗CD3单抗刺激人T细胞活化。结论重组蛋白B7 2 (IgV +C)在第一信号存在下可活化T细胞 。
Objective To clone, express and investigate the costimulatory activity of B7-2(IgV+C), extracellular domain of B7-2(CD86). Methods B7-2(IgV+C) gene was amplified from B7-2 cDNA by PCR and then cloned into an prokaryotic expression vector pGEX-4T-3. The engineered E.coli were used to express protein. After denaturation and renaturation, the protein was used to stimulate human T lymphocytes cooperated with anti-CD3 in vitro. 3H-TdR incorporation was used to detect the activation of T cells. Results The plasmid pGEX-4T-3/hB7-2(IgV+C) was reconstructed and expressed a M r 55×10 3 fusion protein in E.coli at a level of 33% of the total cellular protein. The experiment in vitro suggested that the fusion protein activated human T lymphocyte. Conclusion With the first signal, the recombinant protein B7-2(IgV+C) had costimulatory avtivity to activate T cells.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2002年第2期131-133,共3页
Chinese Journal of Microbiology and Immunology
基金
国家自然科学基金资助项目 ( 39470 2 93)
