肺癌患者铜绿假单胞菌感染快速检测方法的应用

Gold immunochromatographic assay for the on-site detection of pseudomonas aeruginosa infection in advanced lung cancer patients

探讨胶体金免疫层析方法在感染铜绿假单胞菌肺癌患者快速床边检验的应用价值。方法:用分子生物学方法克隆OprF基因,原核系统诱导表达并纯化OprF,免疫BALB/c小鼠后通过杂交瘤技术制备特异性的mAb。采用双抗体夹心法,建立了检测铜绿假单胞菌的胶体金免疫层析法,并对该方法的特异性、灵敏度和稳定性进行了评价,对121例晚期肺癌患者临床标本进行了检测。结果:成功克隆OprF基因,经原核诱导表达获得铜绿假单胞菌的OprF。通过杂交瘤技术筛选,经鉴定单克隆抗体3C3B5可作为捕获抗体,多克隆抗体被HRP标记后可作为检测抗体。以传统细菌培养和鉴定方法为参照,用胶体金免疫层析试纸条检测的敏感度为83.3%(45/54),特异性为92.5%(62/67),总符合率为88.4%。结论:成功制备了铜绿假单胞菌胶体金免疫层析法试纸条,实现了晚期肺癌感染患者床边检验和病情检测。中华肿瘤防治杂志,2012,19(2)

To develop a rapid one-step gold immunochromatographic assay to detect pseudomonas aeruginosa infection in terminal lung cancer patients.METHODS： The OprF gene was constructed into the prokaryotic expression vector pET28b.The OprF protein was expressed in prokaryotic system.Anti-OprF monoclonal antibody（mAb） was generated by hybridoma technique.To develop gold immunochromatography assay by using mAb,polyclonal antibody（pAb） and colloidal gold labeling technique,based on double antibody sandwich immunoassay.To assess the reletively sensitivity and specificity of gold immunochromatography assay by detecting 121 clinical samples from advanced lung cancer patients.RESULTS： The OprF gene and protein was expressed successfully.The sandwich gold immunochromatography was developed with mAb as detection antibody and pAb as capture antibody.The sersitivity of this method was 83.3%（45/54）,the specificity was 92.5%（62/67）,and the coincidence was 88.4%.Compared with traditional culture method,the gold immunochromatography was damostrated to be good consistency with culture method.CONCLUSION： The rapid one-step gold immunochromatography developed in this study has been proved to be senentity,specific and simple for detection pseudomonas aeruginosa infection in advanced lung cancer patients.