电融合产生黄瓜花叶病毒的单克隆抗体

CMV SPECIFIC MONOCLONAL ANTIBODIES PRODUCTION BY ELECTROFUSION

用提纯的黄瓜花叶病毒(CMV)种传SS-30株和豇豆株(P株)混合免疫BALB/c小鼠,取其脾细胞与小鼠骨髓瘤细胞SP2/0,用BAEKON 2000基因转移仪使其融合。通过2—3次简易、快速的有限稀释,获得6个能持续分泌抗体的杂交瘤细胞株,所获抗体分属IgG和IgG_(2a),并能诱导高滴度腹水抗体。间接ELISA试验证明这些单克隆抗体对种传CMVSS-30株、P株和非种传CMV的黄化、日本、山东、香蕉、向日葵和Q等分离物有特异性反应,与C_((?))分离物的反应性较弱,而对花生矮化病毒(PSV)无交叉反应。利用间接ELISA测定了几个单克隆抗体的亲和力常数。单克隆抗体C_7H_3、A_6H_4与G_4G_3、G_4H_3所抗的抗原决定簇相距较远,而C_7H_3与A_6H_4,G_4G_8与G_4H_5为抗同一抗原决定簇的单克隆抗体。

Six hybridomas stably producing high titer monoclonal antibodies against CMV have been derived from electrofusion the spleen cells of CMV-ss-30 and CMV-p immunized BALB/c mice with SP2/0 using BAEKON 2000 Advanced Gene Transfer System. Then they were cloned by 2—3 times of simple, efficient, standard-dilution. The monoclonal antibodies were of IgG_1 and IgG_2. subclass. Their specificities were determined by indirect ELISA and show quite strong binding ability against ss-30, p, yellow, Japan, Shandong, banana, sunflower and Q isolates of CMV. The lower binding ability was observed with C_(59) isolate of CMV. They were not reacted at all with PSV. The affinity cosrant and additivity index of monoclonal antibodies have been determined by indirect ELISA. The C_7H_3 and A_6H_4, G_4G_8 and G_4H_5 have same determinant. The C_7H_3, A_6H_4 and G_4G_8, G_4H_5 might recognized different determinant.