全自动核酸提取平台高敏HBV DNA 定量检测的性能评价

Evaluation of an automatic nucleic acid extraction platform for high sensitive HBV DNA quantitative detection

目的验证Natch S全自动核酸提取平台高敏HBV DNA定量检测的性能。方法参考美国临床实验室标准化协会批准指南(CLSI-EP),采用Natch S全自动核酸提取平台及实时荧光定量聚合酶链反应分析系统检测高敏HBV DNA,评价该方法的精密度、正确度、线性范围、检测能力和抗污染能力等性能指标,采用化学发光微粒子免疫检测法检测HBV标志物和IFCC速率法检测丙氨酸氨基转移酶(ALT),并分析两者之间的相互关系。结果精密度方面,高(10~7 IU/ml)、中(10~3 IU/ml)和低(10~2 IU/ml)浓度样本的批内与批间精密度CV值均≤5.0%;正确度方面,参加2017-2018年全国临床检验中心室间质评正确率100%,且实测值与靶值偏倚均<±0.20 lg IU/ml;线性范围方面,在(1.0×10~2~5.0×10~9)IU/ml范围内有良好的线性关系(Y=0.4+0.97X,R^2=0.999≥0.98,P<0.05);检测能力方面,检出限30 IU/ml和定量限100 IU/ml的检出率100%,且定量限100 IU/ml的CV值为3.21%;抗污染能力方面,与高浓度(>10~6 IU/ml)阳性样本间隔放置的阴性样本均未检出阳性结果;临床应用评价,566例慢性乙型肝炎(乙肝)患者血清样本高敏HBV DNA载量分布情况以<30 IU/ml组为主,占45.94%,与其他组ALT异常构成比差异有统计学意义(P均≤0.043),同一HBV DNA载量水平HBeAg>1.0 S/Co组与HBeAg≤1.0 S/Co组ALT异常构成比差异无统计学意义(P均≥0.61)。结论基于Natch S全自动核酸提取平台的高敏HBV DNA定量检测各项分析性能指标均表现良好,符合PCR检测要求,且对低病毒载量慢性乙肝患者抗病毒治疗监测具有重要意义。

Objective To evaluate the performance of the Natch S automatic nucleic acid extraction platform for high-sensitivity HBV DNA quantitative detection.Methods Based on the American standard CLSI-EP,the Natch S automatic nucleic acid extraction platform and PCR analysis system was applied to detect the high-sensitivity HBV DNA.The precision,accuracy,linear range,detection ability and anti-pollution ability of this method was analyzed as well as the correlation with HBV DNA,ALT and HBeAg.Results Both the intra-assay and inter-assay precision CV values were≤5.0%.The correct rate of EQA was 100%from 2017 to 2018.The linear range was good in the range of(1.0×10~2-5.0×10~9)IU/ml(Y=0.4+0.97X,R^2=0.999≥0.98,P<0.05).The detection rate of 30 IU/ml LOD and 100 IU/ml LOQ was 100%,and the CV value of 100 IU/ml LOQ was 3.21%;No positive results were detected in the negative samples in terms of anti-pollution ability.The distribution of HBV DNA in 566 patients with chronic hepatitis B was mainly in the group of<30 IU/ml(45.94%).There was a statistically significant difference in the ratio of ALT abnormalities between the<30 IU/ml group and other groups(P≤0.043).There was no significant difference in the ratio of ALT abnormalities between HBeAg>1.0 S/Co group and the HBeAg≤1.0 S/Co group at the same HBV DNA level(P≥0.61).Conclusions Based on the Natch S automatic nucleic acid extraction platform,the high-sensitivity HBV DNA quantitative detection performance indicators are all good for monitoring of anti-viral treatment of chronic hepatitis B patients with low levels of HBV DNA.