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目的制备抗人二羰基/L-木酮糖还原酶(Dicarbon-yl/L-xylulosereductase,DCXR)的单克隆抗体(mAb)并进行初步鉴定。方法将正常成人肝组织匀浆离心并分离线粒体,用线粒体总蛋白免疫BALB/c小鼠,采用杂交瘤技术制备mAb,并通过间接ELISA法、Westernblot及免疫组化的方法对mAb进行特性鉴定,通过免疫沉淀联合质谱、Uni-ZAPXR表达文库筛选鉴定抗原。结果获得1株可稳定分泌抗人DCXRmAb的杂交瘤细胞系。mAb的Ig亚类(型)为IgG1(κ),该mAb可用于ELISA检测、Westernblot、免疫组化和免疫沉淀实验。结论成功制备了抗人DCXR的mAb,为DCXR的研究提供了有力的工具。
Objective To prepare monoclonal antibodies against human dicarbonyl / L-xylose reductase (DCXR) and to identify them. Methods Normal adult liver homogenates were centrifuged and isolated from mitochondria. BALB / c mice were immunized with total mitochondrial protein. The mAbs were prepared by hybridoma technique. The characteristics of mAbs were identified by indirect ELISA, Western blot and immunohistochemistry. Uni-ZAPXR expression library was screened for antigen identification by immunoprecipitation combined with mass spectrometry. As a result, a hybridoma cell line stably secreting anti-human DCXRmAb was obtained. The mAb has an Ig subclass (type) of IgG1 (κ) that can be used in ELISA assays, Westernblot, immunohistochemistry and immunoprecipitation assays. Conclusion The mAb against human DCXR was successfully prepared, which provided a powerful tool for the study of DCXR.