粪肠球菌核酸检测试剂国家参考品的研制  

Development of national reference materials for Enterococcus faecallis nucleic acid detection kit

作　　者：王春娥[1] 石继春[1] 徐潇[1] 刘茹凤[1] 杜宗利 龙新星 徐颖华[1] 叶强[1] WANG Chun-E;SHI Ji-Chun;XU Xiao;LIU Ru-Feng;DU Zong-Li;LONG Xin-Xing;XU Ying-Hua;YE Qiang(Key Laboratory of the Ministry of Health for Research on Quality and Standardization of Biotech Products,National Institute for Food and Drug Control,Beijing 102629,China)

机构地区：[1]中国食品药品检定研究院,中国医学细菌保藏管理中心,卫生部生物技术产品检定方法及其标准化重点实验室,北京102629

出　　处：《食品安全质量检测学报》2021年第12期4801-4806,共6页Journal of Food Safety and Quality

基　　金：国家重点研发计划项目(2018YFC1603900);国家科技重大专项(2018ZX10102-001);国家科技基础条件平台项目(NMRC-2020-2)。

摘　　要：目的研制粪肠球菌核酸检测试剂的国家参考品。方法将9株粪肠球菌和8株非粪肠球菌参考品候选菌株分别在各自适宜的培养条件下培养,收集新鲜培养物进行计数、灭活、稀释和分装,并对参考品进行均匀性和稳定性评估。组织3家实验室对研制的参考品进行了协作标定。结果研制了由9份阳性候选参考品、8份阴性候选参考品、重复性参考品和最低检出限参考品组成的粪肠球菌参考品。阳性参考品、最低检出限参考品和重复性参考品循环阈值(cycle threshold, Ct)的变异系数(coefficient of variation, CV)均在5.0%以内,分装均匀性良好;参考品各成分的稳定性良好(P值均小于0.05);协作标定结果显示阳性参考品符合率及阴性参考品符合率均为100%,重复性参考品检测Ct值的CV均在5.0%以内,参考品最低检出限为1.0×10^(3)个/mL。结论研制的参考品可用于粪肠球菌核酸检测试剂的质量评价。Objective To develop national reference materials for Enterococcus faecalis nucleic acid detection kit. Methods Nine strains of Enterococcus faecalis and 8 candidate strains of non-Enterococcus faecalis reference samples were cultured under their respective suitable culture conditions. Fresh cultures were collected for counting, inactivation, dilution and packing, and the reference samples were evaluated for homogeneity and stability. Three laboratories were organized to calibrate the developed reference. Results The coefficient of variation(CV) of cycle threshold(Ct) of positive reference, minimum detection limit reference and repeatable reference were all less than 5.0%, and the distribution uniformity was good. The stability of the reference components was good(P<0.05);the cooperative calibration results showed that the coincidence rate of positive reference and negative reference were100%, the Ct value of repeatable reference was less than 5.0%, and the minimum limit of detection reference was 1×10^(3)/mL. An Enterococcus faecalis reference product was prepared, which consisted of 9 positive candidate reference products, 8 negative candidate reference products, repeatable reference products and minimum detection limit reference products. Conclusion The developed reference material can be used for the quality evaluation of nucleic acid detection reagent for Enterococcus faecalis.

关 键 词：粪肠球菌 核酸 国家参考品 均匀性 稳定性 协作标定 

分 类 号：R155.5[医药卫生—营养与食品卫生学]