分泌抗人精子单克隆抗体杂交瘤细胞株的建立及鉴定

建立两株分泌抗人精子单克隆抗体的杂交瘤细胞株G_8与E_(10),采用洗涤人精子免疫Balb/c小鼠,取免疫脾与Balb/c小鼠的Sp2/0骨髓瘤细胞融合,杂交瘤细胞诱生腹水的方法。Elisa,IIF或酶标抗体免疫组化法结果发现,G_8与E_(10)单抗与人精子(射出精子、睾丸、副睾管及输精管中的精子)及睾丸各级生殖细胞均发生反应;与人甲状腺上皮细胞、肾小管上皮细胞、胰腺导管上皮细胞以及胰岛均发生不同程度的交叉反应,与其他33种人体正常组织、9种人体液与分泌液、8种不同种族的动物精子均不发生交叉反应,说明在一般情况下这两种抗体可用于鉴定人类的精液斑以及精液与阴道分泌液的混合斑。两种单抗与人精子的顶体部、赤道部、后核帽、颈部及中段结合,证明精子特异性表面抗原分布于这几个区域。两种抗体均属IgG_1,G_8与E_(10)抗体的效价分别为1:512与1:1024。

ESTABLISHMENT OF HYBRIDOMA CELL LINES (G8,E10) SECRETING MONOCLONAL ANTIBODIES AGAINST HUMAN SPERMATOZOA

Two hybridoma cell lines (G8,E10) secreting monoclonal antibodies against human spermatozoa were established by fusing Sp2/0 mouse myeloma cells with splenocytes from Balb/c mice immunized with washed human spermatozoa. The results of the IIF, Elisa or peroxidase conjugated antibody im- munohistochemical staining demonstrated that both antibodies reacted with the spermatozoa in the human semen, testis, epididymus and vas deferens as well as the germ cells in the testis, cross reacted with the epithelial cells Of the thyroid gland, renal tubules, pancreatic ducts as well as the ilets of Langhans of the pancrease and not cross reacted with 33 normal human tissue cells, 9 different sources of body fluids and secretions as well as 8 different species of animal's spermatozoa. It is concluded that both monoclonal antibodies can be used for the species identification of semen stain in sex assault cases. The results of IIF demonstrated that each antibody bind to the localized region of the sperm cell surface. The antibody binding patterns were restricted to the acromosal cap, equatorial segment, postacromosal region, neck and the midpiece. The observed antibody binding patterns suggest that the human sperm surface is divided into a minimum of five domains. Both monoclonal antibodies were belong to IgG_1 subclass. The titre of Gs was 1∶512, while of E_(10) 1∶1024.