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抗磷脂酰肌醇蛋白聚糖-3单克隆抗体的制备与鉴定

Preparation and characterization of monoclonal antibody against glypican-3.

作者信息

Ma Rui-juan, Wang Sui-hai, Qin Shi-ni, Wang Xiao-bo, Li Gao-fei, Li Ming, Xu Wei-wen

机构信息

School of Biotechnology, Southern Medical University, Guangzhou, P.R. China.

出版信息

Hybridoma (Larchmt). 2012 Dec;31(6):455-61. doi: 10.1089/hyb.2012.0030.

Abstract

Glypican-3 (GPC3) has been reported as a novel serum and histochemical marker for hepatocellular carcinoma (HCC) by several groups. As an oncofetal protein, it is expressed abundantly in the fetal liver, inactive in the normal adult liver, and frequently reactivated in HCC. Immunology reagents are urgently needed to proceed with mechanism-related research, clinical validation, and application. In this report, monoclonal antibodies (MAbs) against GPC3 were made from hyperimmune BALB/c mice by injecting 100 μg of purified antigen intraperitoneally. Hybridomas were screened by indirect enzyme-linked immunosorbent assay (ELISA) using purified protein. Finally 13 mouse hybridomas producing MAbs to GPC3 were established. The MAbs obtained were fully characterized using Western blot analysis, immunofluorescence, and immunohistochemistry. The results showed that these antibodies could be used for preliminary application of the next step mechanism-related research and GPC3 expression level analysis.

摘要

几组研究报告称,磷脂酰肌醇蛋白聚糖-3(GPC3)是一种用于肝细胞癌(HCC)的新型血清和组织化学标志物。作为一种癌胚蛋白,它在胎儿肝脏中大量表达,在正常成体肝脏中不活跃,而在HCC中经常重新激活。开展与机制相关的研究、临床验证及应用迫切需要免疫试剂。在本报告中,通过向超免疫BALB/c小鼠腹腔内注射100μg纯化抗原制备了抗GPC3单克隆抗体(MAb)。使用纯化蛋白通过间接酶联免疫吸附测定(ELISA)筛选杂交瘤。最终建立了13株产生抗GPC3 MAb的小鼠杂交瘤。使用蛋白质印迹分析、免疫荧光和免疫组织化学对获得的MAb进行了全面表征。结果表明,这些抗体可用于下一步与机制相关研究及GPC3表达水平分析的初步应用。

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