Qian Li, Qian Guan-Xiang
Research Centre of Molecular Biology, Shanghai Second Medical University, Shanghai 200025, China.
Sheng Wu Hua Xue Yu Sheng Wu Wu Li Xue Bao (Shanghai). 2002 Sep;34(5):547-52.
To study the effect of OVA-linker-beta2m fusion protein as an immunogen to induce CTL mediated immune response in mice. The beta2m gene was amplified by PCR. OVA(257-264) peptide sequence and an 8-amino acid linker were added to the COOH-terminus of beta2m gene, and then cloned into pE42b(+) vector. After that, gene expression, protein purification and refolding were conducted. Then the protein product was used as an immunogen to induce an efficient CTL mediated immune response in vivo. Specific CTL responses were demonstrated by H-2K(b)-OVA tetramer staining and cytotoxicity assay. Extracelluar IFN-gamma was also quantitatively analyzed. The results showed that specific CTL response could be induced in vivo by OVA-linker-beta2m fusion protein. Generation of terameric peptide-MHC complexes in vitro is a powerful tool to stain specific CTL.
研究卵清蛋白（OVA）-连接肽-β2微球蛋白（β2m）融合蛋白作为免疫原诱导小鼠体内细胞毒性T淋巴细胞（CTL）介导的免疫反应的效果。通过聚合酶链反应（PCR）扩增β2m基因。将OVA（257-264）肽序列和一个8氨基酸连接肽添加到β2m基因的羧基末端，然后克隆到pE42b（+）载体中。之后，进行基因表达、蛋白质纯化和复性。然后将蛋白质产物用作免疫原在体内诱导高效的CTL介导的免疫反应。通过H-2K（b）-OVA四聚体染色和细胞毒性试验证明特异性CTL反应。还对细胞外γ干扰素进行了定量分析。结果表明，OVA-连接肽-β2m融合蛋白可在体内诱导特异性CTL反应。体外生成三聚体肽-主要组织相容性复合体（MHC）是标记特异性CTL的有力工具。
