家蚕微孢子虫单克隆抗体的制备及鉴定

用家蚕微孢子虫(Nosema bombycis)总蛋白为抗原,采用长程免疫法免疫BALB/c小鼠,取4次免疫后的脾细胞与SP2/0骨髓瘤细胞用PEG-1500进行细胞融合,第1次融合率为13.3%,第2次融合率为55.4%。建立间接酶联免疫吸附测定法(enzyme-linked immunosorbent assay,ELISA),用于检测杂交瘤细胞上清中的单克隆抗体。初检阳性率分别为4.68%和5.26%。经3次亚克隆筛选获得了2株能稳定分泌抗家蚕微孢子虫的单克隆抗体的杂交瘤细胞株,命名为2G10和2B10。间接免疫荧光试验(indirect immunofluorescence test,IFAT)和Western-bloting分析证实所获得的单克隆抗体是特异针对家蚕微孢子虫蛋白的,将在孢子孢壁蛋白的研究中发挥重要作用。

Preparation and Identification of Monoclonal Antibodies Against Nosema bombycis

The BALB/c mice were immunized four times with the total protein of Nosema bombycis,then the spleen cells of the hyperimmunized mouse and SP2/0 cells were fused with PEG-1500.The cell fusion rate was 13.3% at the first time and 55.4% at the second time.An indirect enzyme-linked immunosorbent assay(ELISA) was developed to detect monoclonal antibodies (McAb) secreted by hybridoma cell lines.And the positive rate was 4.68% and 5.26% at the first time and the second time,respectively.Two hybridoma cell lines secreting monoclonal antibodies,named 2G10 and 2B10,were developed by three times of subclone.The specificity of McAb against Nosema bombycis' proteins was identified by means of indirect immunofluorescence test(IFAT) and Western-blotting.The McAb should play a vital role in researching spore wall proteins of N.bombycis.