百日咳杆菌核酸检测试剂国家参考品的研制  

Development of national reference panel for detection kit for Bordetella pertussis nucleic acids

作　　者：夏德菊 周海卫[1] 王薇 许四宏[1] XIA De-ju;ZHOU Hai-wei;WANG Wei;XU Si-hong(Division II of Diagnostic Kits for Infectious Diseases,National Institutes of Food and Drug Control,Beijing 100050,China)

机构地区：[1]中国食品药品检定研究院传染病诊断试剂二室,北京100050

出　　处：《中国生物制品学杂志》2021年第12期1449-1455,1462,共8页Chinese Journal of Biologicals

基　　金：国家突发急性传染病诊断试剂参考品及评价技术研究(2018ZX10102001)。

摘　　要：目的研制百日咳杆菌核酸检测试剂国家参考品,确定其质量标准。方法通过对21份病原体培养物的复核、确证,关键抗原位点等位基因分析、红霉素耐药位点测序分析,组成百日咳杆菌核酸检测试剂国家参考品,经多家实验室协助标定,确定参考品的质量标准,并对参考品进行均匀性及稳定性评估。结果百日咳杆菌核酸检测试剂国家参考品由12份阴性、10份阳性、1份检出限和1份精密度样品组成。质量标准为12份阴性参考品应均检出阴性;10份阳性参考品应均检出阳性,若为百日咳杆菌红霉素耐药位点检测试剂盒,P1、P3、P6、P7和P9需检出耐药位点阳性;精密度参考品重复检测10次,要求均为百日咳杆菌阳性,且其检测值的变异系数(CV)应不大于5.0%,若为百日咳杆菌红霉素耐药位点检测试剂盒,重复检测10次,要求均为耐药位点阳性,且其检测值的CV不大于5.0%;最低检出限参考品要求对于选择IS481靶区的试剂,浓度为100cfu/mL及以上时,检测结果应为百日咳杆菌阳性;其他靶区试剂要求浓度为1×10^(4)cfu/mL及以上时,检测结果应为百日咳杆菌阳性,若为百日咳杆菌红霉素耐药位点检测试剂盒,要求浓度为1×10^(4)cfu/mL及以上时,需检出耐药位点阳性。参考品的均匀性及稳定性均符合要求。结论研制的百日咳杆菌核酸检测试剂国家参考品可用于百日咳杆菌核酸检测试剂的质量控制及评价。Objective To develop a national reference panel for detection kit for Bordetella pertussis and establish its quality standard.Methods By screening of 21 pathogen cultures,analysis of alleles at key antigen sites and sequencing analysis of erythromycin resistance sites,the candidate reference was composed and calibrated by several laboratories to determine the quality standard,and evaluated for uniformity and stability.Results The national reference panel for detection kit for B.pertussis consisted of 12 negative,10 positive,one limit of detection and one precision samples.The quality standard was as follows:all the test results of 12 negative references were negative;all the test results of 10 positive references were positive;if a detection kit for erythromycin resistance sites of B.pertussis was used,positive resistance sites should be found in positive references P1,P3,P6,P7 and P9;the precision reference should be tested for 10 times repeatedly,and all the results should be positive with a CV of not more than 5.0%and,if a detection kit for erythromycin resistance sites in B.pertussis was used,the test should be repeated for 10 times,and all the results should be positive with a CV of not more than 5.0%;for the kits in which the IS481 target area was selected,the test result by the reference for limit of detection should be positive for B.pertussis when the concentration was not less than 100 cfu/mL;for the kits in which other target areas were selected,test result should be positive when the concentration was not less than 1×10^(4)cfu/mL;however,if a detection kit for erythromycin resistance sites was used,positive resistance sites should be found when the concentration was not less than 1×10^(4)cfu/mL.Both the uniformity and stability of the reference met the relevant requirements.Conclusion The developed national reference panel may be used for the quality control and evaluation of detection kit for B.pertussis nucleic acids.

关 键 词：百日咳杆菌 核酸检测试剂 红霉素耐药 国家参考品 

分 类 号：R447[医药卫生—诊断学]