基于化学发光磁酶免疫分析技术检测单增李斯特菌  

Detection of Listeria monocytogenes through Chemiluminescent Magnetic Enzyme Immunoassay

作　　者：范龙兴[1,2] 宁保安[2] 孙智勇[3] 白家磊[2] 彭媛[2] 曲晓辰 刘超[2] 乌恩琦[1] 高志贤[2] 刘颖[1] 

机构地区：[1]内蒙古医科大学公共卫生学院,内蒙古呼和浩特010110 [2]军事医学科学院卫生学环境医学研究所天津市环境与食品安全风险监控技术重点实验室,天津300050 [3]中国人民解放军第十一医院,新疆伊宁835000

出　　处：《食品研究与开发》2017年第3期124-129,共6页Food Research and Development

基　　金：国家重大科学仪器设备开发专项(2013YQ14037106)

摘　　要：探索化学发光磁酶免疫分析技术在单核细胞增生性李斯特菌检测中的应用,并初步建立和优化检测方法及条件。将抗LM兔多抗与磁性微球偶联构建免疫磁分离原件,应用双抗夹心法ELISA检测LM全菌抗原,应用化学发光法进行定量检测,应用正交设计优化主要反应条件,建立标准曲线,测试灵敏性与特异性。通过正交试验优化的反应条件为37℃条件下,免疫磁珠用量20μL、含0.1%BSA的PBS封闭15 min,捕获抗原2 h,HRP标记的抗LM单抗稀释至1∶10 000(体积比)特异性结合反应30 min后进行化学发光检测,标准曲线拟合优度0.954,最低检出限10~4 CFU/m L,特异性良好,检测时限为3 h^4 h。To explore the application of chemiluminescent magnetic enzyme immunoassay in the detection of Listeria monocytogenes,and to establish and optimize the detection methods and conditions. The magnetic beads were coupled with rabbit anti LM polyclonal antibody to structure the original of immunomagnetic separation. L. monocytogenes was detected by chemiluminescence through double antibody sandwich ELISA. Orthogonal design was used to optimize the reaction conditions,and to establish the standard curve,the sensitivity and specificity were tested. Through orthogonal design,the optimization of reaction conditions were in 37 ℃,immunomagnetic bead dosage of 20 μL,containing 0.1 % BSA in PBS blocking 15 min,antigen captured 2 h and HRP labeled anti LM monoclonal antibody dilution to 1 ∶ 10 000（体积比）specificity binding reaction 30 min after chemiluminescence detection. The goodness of fit of standard curve is 0.954,the minimum detection limit is 10～4CFU/m L,the specificity is good,and the detection time is 3 h-4 h.

关 键 词：化学发光 免疫磁珠 单增李斯特菌 

分 类 号：O657.3[理学—分析化学] TS207.4[理学—化学]