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目的 利用噬菌体抗体库技术制备抗甲状旁腺激素相关蛋白 (PTHrP)Fab抗体。方法从经PTHrP免疫的Balb/C小鼠脾细胞提取总RNA ,用逆转录 聚合酶链反应 (RT -PCR)扩增一组Ⅰg轻链和重链Fab段基因 ,插入噬菌体载体pComb3,构建了抗PTHrPFab抗体基因库。用合成短肽和重组PTHrP对抗体库进行了富集筛选 ,获得鼠源抗PTHrPFab段基因后 ,进一步在大肠埃希菌表达 ,并对表达产物的特异性作了鉴定。结果 成功构建了抗PTHrP抗体基因库 ,且筛选到PTHrP特异的鼠源单抗Fab段基因。结论 所获得的Fab抗体能特异性地识别PTHrP抗原。
Objective To prepare anti-parathyroid hormone-related protein (PTHrP) Fab antibody using phage antibody library. Methods Total RNA was extracted from spleen cells of Balb / C mice immunized with PTHrP. A group of Ig light chain and heavy chain Fab gene was amplified by reverse transcription-polymerase chain reaction (RT-PCR) and inserted into the phage vector pComb3 Anti-PTHrPFab Antibody Gene Bank. Antibody library was enriched and screened with synthetic short peptide and recombinant PTHrP. After the mouse anti-PTHrPFab gene was obtained, it was further expressed in Escherichia coli and the specificity of the expressed product was identified. Results The gene library of anti-PTHrP antibody was successfully constructed and the PTHrP-specific murine monoclonal antibody Fab fragment gene was screened. Conclusion The Fab antibody obtained can specifically recognize PTHrP antigen.