合成多肽为半抗原制备抗SCF受体单克隆抗体

Generation of mAbs against synthetic SCF receptor peptides

本文成功地建立了以合成的小分子肽与蛋白质交联的偶联物为免疫原，制备单克隆抗体（ｍＡｂ）的技术路线。选定ＳＣＦ受体特异性肽段进行人工合成，将其偶联于ＢＳＡ免疫动物，应用细胞融合技术制备了抗两肽段的杂交瘤细胞７株。ＥＬＩＳＡ法和免疫印迹结果显示，其分泌的ｍＡｂ特异性强，与偶联载体及多种多肽生长因子均无交叉反应。经ＡＰＡＡＰ法鉴定，抗胞内羧基端肽的ｍＡｂＳＢ３０４与ＳＣＦ受体表达细胞系ＨＥＬ有弱的阳性反应；免疫组化测定与胎儿皮肤基底层色素细胞及胃粘膜上皮细胞呈阳性反应，提示ｍＡｂＳＢ３０４可识别天然状态的ＳＣＦ受体。

In order to generate monoclonal antibody to human stem cell factor receptor(SCF R), we synthesized two peptides according to the protein sequence of SCF R. The peptides were linked with carrier protein and then were used as fusion antigens for immunization. Two peptides SCFR 1 and SCFR 2, which correspond to the carboxyl terminal and extracelular portion of SCF R separately, were designed on the basis of computer assisted analysis of the structural features such as antigenicity and homology etc. After immunization and cell fusion process, we have obtained 7 strains of hybridomas producing monoclonal antibodies to these peptides. These antibodies react specifically wth the synthetic peptides but not the carrier protein and other cytokines. In APAAP test with mAb SB304(to SCFR 1), mild positive staining signals were observed in HEL cell line which expresses SCF R. In addition, SB304 can react with epithelial cells and pigmental cells from fetal skin and stomach mucosa by immunohistochemical identification. These results imply that the SB304 can recognize and bind the natural SCF R protein. The subclasses of the 7 strains mAbs include mice IgG1, IgG2 and IgG2b.