呼吸道合胞病毒核酸分型检测性能验证

Performance validation of respiratory syncytial virus nucleic acid typing kit

目的评价呼吸道合胞病毒(RSV)核酸分型检测试剂盒的分析性能。方法留取呼吸道合胞病毒抗原检测阳性和阴性的咽拭子,从咽拭子洗脱液提取核酸,用RSV RNA分型检测试剂盒检测。核酸检测的扩增产物送测序公司测序,测序结果与NCBI标准序列比对确认为RSV核酸并分型,测序结果与待评价试剂盒检测结果比对评价正确度。取RSV核酸检测弱阳性的样本,每天重复检测4次,连续检测5 d,以检测Ct值评价检测精密度。将RSV A型和B型的RNA标准品稀释到检测限,验证检测下限。取微生物室咽拭子培养常见病原菌的纯培养菌液,验证检测试剂的交叉无反应性。结果通过实验验证检测试剂盒的符合率为100%,批内、批间精密度(CV)≤5%,检测下限为100 copy/mL,与常见同部位病原菌无交叉反应。结论RSV核酸检测试剂和仪器组成的检测系统各项质量指标符合试剂说明书的质量参数,可开展临床标本的检测,为临床早诊断、早治疗提供重要依据。

Objective To evaluate the analytical performance of respiratory syncytial virus(RSV)nucleic acid typing test kit.Methods The positive and negative pharyngeal swabs for RSV antigen were retained.The nucleic was extracted from pharyngeal swabs elution and detected by using the RSV RNA genotyping kit.The amplified products of nucleic acid detection were sent to the sequencing company for sequencing.The sequencing results were identified as RSV nucleic acid and typed by the alignment with NCBI standard sequence.The sequencing results were compared with the detected results of the kit to be evaluated for evaluating the accuracy.The weakly positive RSV nucleic acid samples were repeatedly tested for 4 times a day during consecutive 5 d.The detection precision was evaluated by Ct value.The RSV RNA standards of type A and B was diluted to the detection limit for verifying the lower limit of detection.The pure culture liquid of common pathogenic bacteria in pharyngeal swab was used to verify the cross-reactivity of detection reagents.Results The coincidence rate of the test kit was 100%.The precision of in-batch and inter-batch detection were≤5%.The detection lower limit of the kit was 100 copy/mL,which had no cross reaction with common pathogens at the same site.Conclusion The various quality indexes of the detection system composed of RSV nucleic acid detection reagent and instrument conform to the quality parameters of reagent instruction,which can be used to conduct the detection of clinical samples to provide the important basis for clinical early diagnosis and early treatment.