摘要
用人工合成沙眼衣原体主要外膜蛋白(MOMP)氨基端1/4肽链免疫8周龄雄性BALB/c小鼠3只,第14天用L1/440/Bu原体加强免疫,第24天将免疫反应良好的1只小鼠脾细胞与NS-1瘤细胞融合。用1/4MOMP和L1原体抗原包被的聚丙乙烯板检测上清中抗体,淘汰与鹦鹉热衣原体种(EAE株)、肺炎衣原体种(ATCCVR1310株)以及正常组织培养细胞(BGMK)有交叉反应的克隆,最后得到4株抗沙眼衣原体特异性单克隆抗体(MAbs),其抗体属IsG1和IgG2a亚类。微量免疫荧光试验(micro-IF)发现4株MAbs均与本实验室制备的沙眼衣原体L1、L2、A、B、C、E原体及L2包涵体抗原发生反应,并与沙眼衣原体所有15个标准血清型结合,其腹水MAbs滴度≥1:12800。免疫印迹试验显示4株MAbs均与沙眼衣原体分子量为40000的MOMP反应。
The synthesized one quarter N-terminal MOMP of C. trachomatis was used for primary immunization of three male BALB/c mice(8 weeks of age), and the boost with C. trachomatis L1/440/Bu elementary bodies(EBs) was followed on day 14. Spleen cells from one mouse with good response of immunization were fused with murine myeloma NS-1 cells on day 24. The hybrid cell suspension was seeded into the wells of 96-well microtest plates which contained macrophage feeder layers. Anti-chlamydial antibodies in culture fluids were screened by ELISA with 1/4 MOMP & L1 EBs coated 96-well trays. Positive wells were cloned by limiting dilution. Four clones which secreted immunoglobulin G1&G2a class were obtained after elimination of those clones that produced antibodies to C. psittaci strain EAE,C. pneumoniae strain ATCC VR1310 and uninfected BGMK cells. In micro-IF test, we found that the all four clones of MAbs reacted with our laboratory prepared L1,L2 ,A,B,C, EEBs, L2 tissue culture inclusions, as well as the EBs of all 15 standard serovars of C. trachomatis. The titers of their ascites were more than 1: 1 2800 in micro-IF test. It was shown that the four clones of MAbs reacted predominantly with 40000 MOMP of C. trachomatis L1 in Western blot.
出处
《中国医学科学院学报》
CAS
CSCD
北大核心
1995年第6期428-433,共6页
Acta Academiae Medicinae Sinicae
基金
WHO资助
关键词
沙眼衣原体
单克隆抗体
外膜蛋白
Chlamydia trachomatis
species-specific monoclonal antibody
major outer membrane protein (MOMP)