摘要
目的对抗人中性粒细胞明胶酶相关脂质运载蛋白(NGAL)单克隆抗体H5识别的抗原表位区域进行鉴定。方法设计4个NGAL截短体,通过PCR扩增截短体DNA片段并克隆到PET42a质粒中表达GST-NGAL截短体融合短肽。以此为抗原鉴定H5识别的抗原表位区域。随后根据生物信息学预测结果,化学合成2段潜在B细胞表位肽,用ELISA和Western blot方法进一步鉴定。结果 H5能与NGAL蛋白分子第20~36位氨基酸结合。结论单克隆抗体H5的抗原表位结合位点存在于成熟NGAL蛋白分子氨基末端第20~36位氨基酸处,其深入研究为建立高效灵敏的NGAL免疫学检测体系和进一步研究NGAL分子抗原表位结构奠定基础。
Objective To identify the epitope region of monoclonal antibody H5 against human neutrophil gelatinase-associated lipoprotein(NGAL).Methods Four NGAL truncated DNA fragments were designed and amplified by PCR and cloned into PET42a plasmid to express GST-NGAL truncated fusion peptide.The antigen epitope region recognized by H5 was identified as antigen.Two potential B cell epitope peptides were synthesized and identified by ELISA and Western blot.Results H5 could bind to the amino acid of the NGAL protein at 20-36 position.Conclusion The antigen epitope binding site of monoclonal antibody H5 exists at at the amino terminal 20-36 amino acid of mature NGAL protein molecule,which lays a foundation for the establishment of a highly sensitive immunoassay system for NGAL and the further study of the epitope structure of NGAL molecule.
作者
何莹
胡川闽
陈安
HE Ying;HU Sichuanmin;CHEN An(Department of Clinical Laboratory,the 986th Hospital of People′s Liberation Army Air Force, Xi′an,Shaanxi 710054,China;Department of Clinical Biochemistry,Army Medical University,Chongqing 400038,China)
出处
《国际检验医学杂志》
CAS
2018年第21期2593-2595,共3页
International Journal of Laboratory Medicine
基金
国家高技术发展研究计划"863"计划资助项目(2011AA02A113)