Wang Jie, Zhang Ying-qian, Zhong Guang-ming, Yu Ping
Department of Immunology, Basic Medical School of Central South University, Changsha 410078, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2010 Oct;30(10):2219-23.
To obtain monoclonal antibodies (mAbs) against Chlamydia trachomatis Tarp protein.
Chlamydia trachomatis serovar D recombinant Tarp fusion protein was cloned and expressed. Balb/c mice were immunized with recombinant Tarp fusion protein, and the spleen cells of the immunized mice were fused with SP2/0 mouse myeloma cells. The hybridoma cell lines secreting mAbs against Tarp protein were screened by an indirect immunofluorescence assay and subcloned by limiting dilution culture. The specificities of these mAbs to Tarp were determined by ELISA, and their isotype and chlamydial species specificity identified by an indirect immunofluorescence assay.
Recombinant GST-Tarp fusion protein with a relative molecular mass of about 136 000 was successfully cloned and expressed. Seven hybridoma cell lines stably secreting specific mAbs against Tarp protein were obtained. All the 7 mAbs reacted strongly with Tarp protein but not with other chlamydial proteins. Two mAbs were identified to belong to IgG2a isotype and the other 5 to IgG1 isotype. All the 7 mAbs reacted strongly with chlamydia serovar A, D, and L2, but not with MoPn, 6BC, or AR39.
The highly specific mAbs against Tarp protein have been obtained to facilitate further study of the structure and function of Chlamydia Tarp protein.
获得抗沙眼衣原体Tarp蛋白的单克隆抗体（mAb）。
克隆并表达沙眼衣原体D血清型重组Tarp融合蛋白。用重组Tarp融合蛋白免疫Balb/c小鼠，将免疫小鼠的脾细胞与SP2/0小鼠骨髓瘤细胞融合。通过间接免疫荧光试验筛选分泌抗Tarp蛋白mAb的杂交瘤细胞系，并通过有限稀释培养进行亚克隆。通过ELISA测定这些mAb对Tarp的特异性，并通过间接免疫荧光试验鉴定其亚型和衣原体种属特异性。
成功克隆并表达了相对分子质量约为136 000的重组GST-Tarp融合蛋白。获得了7个稳定分泌抗Tarp蛋白特异性mAb的杂交瘤细胞系。这7个mAb均与Tarp蛋白强烈反应，但不与其他衣原体蛋白反应。鉴定出2个mAb属于IgG2a亚型，另外5个属于IgG1亚型。这7个mAb均与衣原体A、D和L2血清型强烈反应，但不与MoPn、6BC或AR39反应。
已获得针对Tarp蛋白的高特异性mAb，有助于进一步研究沙眼衣原体Tarp蛋白的结构和功能。
