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目的:鼠抗人CD83功能性单克隆抗体(mAb)的研制及其生物学特性的鉴定。方法:以人CD83转基因细胞L929/CD83为免疫原,常规免疫6～8周龄的雌性BALB/c小鼠;采用B淋巴细胞融合技术,将免疫小鼠脾脏细胞与Sp2/0融合,以L929/CD83、293/CD83转基因细胞为抗体筛选阳性细胞,经免疫荧光标记分析对杂交瘤进行反复筛选和多次的克隆化培养;采用Ig类和亚类快速定性试纸法、染色体核型分析、竞争结合抑制试验、间接免疫荧光法、Westernblot对mAb的生物学特性进行鉴定。结果:获得1株稳定分泌鼠抗人CD83mAb的杂交瘤细胞株(命名为9D8),该mAb能特异性地识别成熟的DC、活化的T细胞及肿瘤细胞株Daudi、8226表达的CD83分子,该mAb识别的位点不同于商品化抗人CD83mAb(HB15e)。结论:成功地研制1株鼠抗人CD83mAb,识别位点与HB15e不同,为更好地研究该分子的功能提供良好的物质基础。
OBJECTIVE: To develop and characterize a murine anti-human CD83 monoclonal antibody (mAb). Methods: Human BALB / c mice of 6-8 weeks old were routinely immunized with human CD83 transgenic L929 / CD83 cells. The spleen cells of immunized mice were fused with Sp2 / 0 by B lymphocyte fusion technique, / CD83, 293 / CD83 transgenic cells were screened by antibody, and the hybridomas were screened repeatedly and cloned by immunofluorescent labeling analysis. Rapid qualitative test strips of Ig and subclasses, chromosome karyotype analysis, competition Combined with inhibition test, indirect immunofluorescence and Western blot, the biological characteristics of mAb were identified. Results: A hybridoma cell line stably secreting mouse anti-human CD83 mAb (named as 9D8) was obtained. This mAb specifically recognizes CD83 molecules expressed by mature DC, activated T cells and tumor cell line Daudi, 8226 The site of mAb recognition differs from the commercial anti-human CD83 mAb (HB15e). CONCLUSION: One mouse anti-human CD83 mAb was successfully developed and its recognition site is different from that of HB15e, providing a good material basis for better studying the function of this molecule.