HBV-HCV-HIV三联荧光PCR检测的内标浓度选择研究

目的选择一个在乙型肝炎病毒-丙型肝炎病毒-人类免疫缺陷病毒(HBV-HCV-HIV)三联荧光PCR检测试验中既可有效监控假阴性结果出现,又对阳性结果影响最小的内标浓度。方法应用不同浓度的内标参入酶链聚合反应(PCR)反应,确定最佳内标参入量;并在适量内标浓度下,检测低浓度标本的阳性检出率。结果由内标浓度5拷贝/PCR、10拷贝/PCR、20拷贝/PCR、50拷贝/PCR、100拷贝/PCR 5个浓度中,优选出最适的内标浓度为20拷贝/PCR,此条件下,阴性标本的内标检出率为100%,检测灵敏度标本的阳性检出率与无内标样本差异无统计学意义。结论合适浓度的内标参与荧光PCR检测能有效地解决了每个标本的质控问题,指示反应体系(试剂耗材)与检测体系(仪器)的有效性。

Study on the optimal concentration of the internal amplification control applied to real-time PCR on HBV-HCV-HIV kits

Objective To determine an optimal concentration of the internal control which can effectively avoid the false negative result and produce a smallest impact on low level positive specimen in HBV-HCV-HIV combined real-time polymerase chain reaction(PCR).Methods The internal amplification control of different concentrations was interpolated to PCR in order to determine the optimal concentration of the internal amplification control;and at the optimal concentration,the positive rate was detected in low concentration level specimen.Results Among five different concentrations of 5,10,20,50 and 100 copies/PCR,the optimal concentration of the internal amplification control was 20 copies/PCR.Under this condition,the internal amplification detection rate of negative specimen was 100%;there was no significant difference in the low level specimen with and without internal amplification control detection rate.Conclusions The appropriate concentration of internal amplification control interpolated real-time PCR detection could effectively monitor each specimen reaction and instruct the validity of the reaction system(reagent and consumed material) and the measure system(instrument).