抗肾综合征出血热病毒人单克隆抗体杂交瘤细胞株的建立及初步鉴定  

Generation of human monoclonal antibodies against hemorrhagic fever with renal sydrome (HFRS) virus

作　　者：赵洪礼[1,2] 张健[1,2] 李忠义[1,2] 张秀梅 姜绍谆[1,2] 刘江秋 韦星呈[1,2] 何亦祥 

机构地区：[1]沈阳军区军事医学研究所 [2]辽宁省基础医学研究所

出　　处：《中华实验和临床病毒学杂志》1997年第1期66-68,共3页Chinese Journal of Experimental and Clinical Virology

摘　　要：以亲和层析技术纯化的肾综合征出血热病毒（ＨＦＲＳＶ）结构蛋白（５５０００，６７０００）为特异性抗原，体外免疫正常人外周血淋巴细胞（ＰＢＬ），然后将体外免疫的淋巴细胞与人－鼠种间杂交瘤细胞（Ｋ６Ｈ６／Ｂ５）融合，经ＥＬＩＳＡ间接法筛选出２株（２Ｄ５，１Ｂ７）分泌抗－ＨＦＲＳＶ人单克隆抗体（Ｈ－ＭｃＡｂ）杂交瘤细胞株，４次克隆化后，１００％阳性。对２Ｄ５株初步鉴定表明，其抗体类型为ＩｇＧ１；培养上清中抗体浓度为２０～３０μｇ／ｍｌ；特异性免疫荧光反应证明２Ｄ５株Ｈ－ＭｃＡｂ是ＨＦＲＳＶ特异性；中和效价为１∶１６０；体外连续传代６个月仍稳定分泌抗体。Peripheral blood lymphocytes (PBL) were obtained from healthy donors and treated with L-leucy-leucin methy (LeuLeu-OMe). The LeuLeu-OMe treated PBL were in vitro immunized with HFRS virus proteins (55kD,67kD) for 6 days, and then fused with K6H6/B5 cells in medium containing 45% (PEG) and 7.5% DMSO. The fused cells were seeded in 96-well plates at cell density 0.5-1×10 5 cells/well in HAT supplemented RPMI-1640 medium. After 14-20 days, 3 positive wells were deteced by ELISA and two (1B7, 2D5) of the 3 hybridoma were stable after 4 time cloning. The hybridoma 2D5 appears to be with high ability of production of the antibodies at a concentration of 20-30μg/ml/10 6 cells/24h. It was proved by IFAT that 2D5 McAb was specific for HFRSV.

关 键 词：肾病综合征 出血热病毒 ELISA 单克隆抗体 

分 类 号：R373.32[医药卫生—病原生物学]