基于自动化提取HCV RNA荧光定量检测系统的性能验证

Validation of the performance of a fluorescence quantitative detection system of HCV RNA based on automated extraction

摘要:

目的 基于Stream SP96 全自动核酸提取仪、ABI7500 实时荧光定量PCR仪对丙型肝炎病毒（HCV）RNA 定量检测系统的性能指标进行验证。方法 依据《临床实验室对商品定量试剂盒分析性能的验证》WS/T 420-2013 性能 验证方案，采用Stream SP96 全自动核酸提取平台及ABI7500 荧光定量PCR仪检测HCV RNA，对其准确度、精密度、线性 区间、检测限、定量限和抗干扰能力等进行方法学性能验证。结果 低、高浓度标准物质的均值与靶值的误差分别为 0.20 和 0.25，均小于靶值对数值±0.4 log；低浓度样本的批内、批间不精密度变异系数值分别为 0.79%、1.01%，高浓度样本的 批内、批间不精密度变异系数值分别为 0.52%、1.22%，均<5%；线性相关系数r>0.980，线性区间可达 20~1.0×10 8 ，呈良好 线性（R 2 =0.997 3）；检出限为 20 IU/mL，最低定量限为 50 IU/mL；含胆红素（300 mg/L）、血红蛋白（300 g/L）、甘油三酯 （3 000 mg/L）的干扰物质对样本检测结果无影响。结论 HCV RNA 实时荧光PCR定量法检测系统准确度、精密度、线 性范围、检出限、定量限、抗干扰能力均符合厂家声明，能够满足临床对HCV RNA定量检测的需求。

Abstract:

Objective To verify the performance indicators of HCV RNA quantitative detection system based on Stream SP96 Automated Nucleic Acid Extractor and ABI 7500 Real-Time PCR System. Methods According to the WS/T 420-2013 Performance Verification Scheme in the Verification of Analytical Performance of Commercial Quantitative Kits by Clinical Laboratory, the Stream SP96 Stream SP96 Automated Nucleic Acid Extractor and ABI 7500 Real-Time PCR System were used to test HCV RNA, and methodological performance verification was performed on the accuracy, precision, linear interval, limit of detection, quantitative limit and anti-interference ability. Results The errors between the mean and target values of the low and high- concentration reference materials were 0.20 and 0.25, respectively, which were less than the logarithmic value of the target value±0.4 log. The intra-batch and inter-batch precision coefficient of variation (CV) values for low-concentration specimens were 0.79% and 1.01%, respectively, and those for high-concentration specimens were 0.52% and 1.22%, respectively, which were all <5%. The linear correlation coefficient r> 0.980, and the linear interval was up to 20-1.0×10 8 , showing good linearity (R 2 =0.997 3). The limit of detection was 20 IU/mL and the lowest limit of quantitation was 50 IU/mL; The interfering substances containing bilirubin (300 mg/L), hemoglobin (300 g/L), and triglyceride (3 000 mg/L) had no effect on the specimen detection results. Conclusion The HCV RNA real-time PCR quantitative testing systems have the accuracy, precision, linear range, limit of detection, limit of quantitation and anti-interference ability in line with the manufacturer’s statement, and they can fully meet the clinical requirements for HCV RNA quantitative testing.