抗人层黏连蛋白单克隆抗体的制备及特性鉴定

Praparation and characterrization of monoclonal antibodies against humen laminin

目的:制备抗人层黏连蛋白(laminin,LN)单克隆抗体(mAb)并鉴定其特性。方法:以人LN免疫BALB/c小鼠,采用杂交瘤技术制备抗人的mAb;同时采用间接ELISA法柃测mAb的腹水效价及mAb的相对亲和力;采用ELISA法鉴定mAb的Ig亚类、进行表位分析及特异性鉴定。结果:获得4株可分泌特异性mAb的杂交瘤细胞2A3、2C6、3G7和4H2,其腹水mAb 的效价为3.6 × 104～2.1×106;4株mAb的Ig亚类为IgG1,轻链均属κ型;相对亲和力2C6在1012以上,2A3、3G7和4H2在106以上;其中2株与1个表位结合,另2株与另外的1个表位结合。结论:成功地制备出抗人LN的mAb,为进一步研究LN在一些疾病中的作用提供了工具。

AIM: To prepare monoclonal antibody ( mAb) against hu-men laminin(LN) and identify their properties. METHODS: BALB/c mice were immunized with humen LN, and then mAb was prepared by hybridoma technique. The titres of mAbs in ascetic fluid and relative affinity of mAb were determined by indirect ELISA test. Ig subgroup, specificity and combine dot of mAb were analysed by ELISA. RESULTS: 4 hybridoma cell lines 2A3, 2C6, 3G7 and 4H2 secreting anti -humen LN mAb were obtained. Their Ig subgroup belonged to IgG1, light chains were type K, The titres of 4 mAbs in ascitic fluid were 3. 6 × 104 ~ 2. 1 × 106. Relative affinity of mAb 2C6 was above 1012 and relative affinity of other 3 mAbs 2A3, 3G7 and 4H2 was 106, 108 and 108. Epitope recognized by 2 mAb(2A3 and 4H2) differened from the that by 2C6 and 3G7 another. CONCLUSION: 4 mAbs against humen LN has been prepared successfully and possess higher liter, specificity and relative affinity. Preparation of anti-human LN mAbs provide a useful regeant for further studying the role of LN in some diseases.