结核分枝杆菌微滴数字PCR检测试剂盒企业参考品的研制  

Development of Enterprise Reference for Mycobacterium Tuberculosis Microdrop Digital PCR Detection Kit

作　　者：王诺 李小红 周雪 李俊花 危宏平[1] Wang Nuo;Li Xiaohong;Zhou Xue;Li Junhua;Wei Hongping(Wuhan Institute of Virology,Chinese Academy of Sciences,Wuhan 430070,China)

机构地区：[1]中国科学院武汉病毒研究所,武汉430070

出　　处：《黑龙江科学》2023年第8期109-112,共4页Heilongjiang Science

摘　　要：为实现结核分枝杆菌复合群核酸检测试剂盒的质量检验,研制了结核分枝杆菌微滴数字PCR检测试剂的企业参考品。将结核分枝杆菌减毒株H 37 Ra、BCG、耻垢分枝杆菌与海分枝杆菌分别在适宜的培养条件下培养,收集新鲜且充分混匀的培养物,计数后灭活、稀释和分装,对参考品进行稳定性评估。使用结核分枝杆菌PCR检测试剂盒进行验证,研制出了一套专用于微滴数字PCR检测试剂的企业参考品,其由2份结核分枝杆菌灭活菌液组成阳性参考品、2份非结核分枝杆菌灭活菌液组成阴性参考品,最低检出量参考品由10^(3)、10^(2)、10^(1)、10^(0)CFU/mL的结核分枝杆菌(灭活H 37 Ra)菌液组成,精密性参考品由10份10^(2)CFU/mL的结核分枝杆菌(灭活H 37 Ra)菌液组成,可稳定保存,用于结核分枝杆菌PCR检测试剂盒(微滴数字PCR法)的质量评价。本参考品具有较好的准确性、特异性及重复性。In order to realize the quality test of mycobacterium tuberculosis complex group nucleic acid detection kit,the enterprise reference of mycobacterium tuberculosis microdrop digital PCR detection reagent is developed.Mycobacterium tuberculosis attenuated strain H 37 Ra,BCG,mycobacterium smegmatis and mycobacterium marinum are cultivated under suitable cultivation condition,fresh and fully mixed culture is collected,inactivated,attenuated and subpackaged after counting,and the stability evaluation of the references is done.Mycobacterium tuberculosis PCR detection kit is used for verification,and a set of enterprise reference reagent for microdrop digital PCR detection is developed.2 samples of inactivated mycobacterium tuberculosis solution constitutes positive reference,and 2 samples of non-inactivated mycobacterium tuberculosis solution constitutes negative reference.The minimum detectable amount reference is composed of 10^(3),10^(2),10^(1),10^(0)CFU/mL mycobacterium tuberculosis(inactivated H 37 Ra)bacterial solution,and the precision reference is composed of 10^(1)0^(2)CFU/mL mycobacterium tuberculosis(inactivated H 37 Ra)bacterial solution,which can be stably preserved.It can be used to evaluate the quality of the mycobacterium tuberculosis PCR kit(microdrop digital PCR method).This reference has good accuracy,specificity and repeatability.

关 键 词：结核分枝杆菌 微滴数字PCR法 企业参考品 

分 类 号：R446.5[医药卫生—诊断学] R52[医药卫生—临床医学]