中国免疫学杂志2025，Vol.41Issue(6)：1484-1489,6.DOI:10.3969/j.issn.1000-484X.2025.06.037
兔抗人FAM21多克隆抗体的制备与鉴定
Preparation and identification of rabbit polyclonal antibody against human FAM21
摘要
Abstract
Objective:To prepare rabbit polyclonal antibody against human FAM21 and analyze antibody specificity.Methods:Using the plasmid encoding human FAM21 full-length gene as a template,the nucleotide sequence of its 2 431～3 006 base was amplified by PCR and connected to the pGEX-6p-1 prokaryotic expression vector to construct pGEX-6P-1-FAM21 recombinant plasmid expressing the 811～1 002 amino acid fragment of FAM21.The recombinant plasmid was transformed into BL21(DE3)compe-tent Escherichia coli and was expressed inductively,and the protein was purified using GST fusion protein purification magnetic beads.The purified GST fusion protein was used as an antigen to immunize New Zealand rabbits,and the collected antiserum was purified by an agarose column containing GST protein.The specificity of antibody was detected by Western blot and immunofluorescence assay in stable FAM21 knockdown HeLa cells.Results:The pGEX-6p-1-FAM21 prokaryotic expression plasmid was successfully constructed and induced to express in BL21(DE3)competent Escherichia coli.The purified GST fusion protein had a molecular weight of approxi-mately 50 kD,and the purified antibody titer from immunized New Zealand rabbits was greater than 1∶128 000,with high specificity.Conclusion:The pGEX-6p-1-FAM21 prokaryotic expression plasmid is successfully constructed,and the rabbit polyclonal antibody against human FAM21 is prepared for Western blot and immunofluorescence assay.关键词
FAM21/原核表达/蛋白表达与纯化/多克隆抗体Key words
FAM21/Prokaryotic expression/Protein expression and purification/Polyclonal antibody分类
医药卫生引用本文复制引用
汤拓,卢彦基,李文龙,王涛,洪鲜,邓志会..兔抗人FAM21多克隆抗体的制备与鉴定[J].中国免疫学杂志,2025,41(6):1484-1489,6.基金项目
黑龙江省自然科学基金优秀青年项目(YQ2022C036) （YQ2022C036）
齐齐哈尔医学院研究生创新基金项目(QYYCX2023-01) （QYYCX2023-01）
齐齐哈尔市科技局联合引导项目(LSFGG-2022045). （LSFGG-2022045）
