血HBV DNA定量检测中的疑难结果分析及其对策

Cause analysis and measures of suspicious results of serum HBV DNA by fluorescence quantitative PCR

目的分析荧光定量PCR检测HBV DNA过程中出现的疑难结果并探讨相应的处理对策。方法采用荧光定量PCR检测血标本HBV DNA含量,分析2008年至2011年本实验室所有送检标本所出现的疑难结果,同时采用两种不同的试剂对疑难结果的标本进行复查;统计两次结果一致性和不一致的比例,并对两次结果不一致原因进行分析。结果 2008年至2011年共有115 154例HBV DNA定量检测血标本,疑难结果的标本数为1 969例,总平均复查率为1.70%;两次结果一致的例数为1 588例(80.65%),两次结果不一致为381例(19.35%);两次结果不一致的原因由试剂因素和试剂外因素造成,试剂因素包括无扩增曲线造成的假阴性和有扩增曲线但扩增效率低下,试剂外因素包括操作污染和其它因素,试剂因素造成两次结果不一致的比例呈逐年上升的趋势。结论采用两种不同的试剂复查疑难结果的标本,大大降低HBV DNA定量的漏检率,避免了因试剂因素和非试剂因素引起的实验误差,提高了检验质量,有效地保证了HBV基因突变标本的HBV DNA准确定量,有利于正确地指导乙肝患者的抗病毒治疗。

Objective To analyze causes and measures of suspicious results of serum HBV DNA by fluorescence quantitative PCR. Methods Blood samples of HBV DNA from 2008 to 2011 were analyzed by fluorescence quantitative PCR.All samples of suspicious results were analyzed and redetected by using two different reagents at the same time.The two results of consistency and inconsistency ratio were evaluated,and the reasons of inconsistent were analyzed.Results There were 115154 cases of HBV DNA and 1969 cases of suspicious results from 2008 to 2011 and the mean ratio of redetection was 1.70%.The cases of two results of consistency and inconsistency were 1588(80.65%) and 381(19.35%),respectively.The two reasons for the inconsistent results included the factors of the reagent,showing no amplification curve caused by the false negative and abnormal low efficiency of amplification curve, and the factors of non reagent,including operating pollution and other factors.The factors of reagent accounting for the inconsistent results in redetection showed a rising trend year by year.Conclusion The way of redetection by using two different reagents for specimens of the suspicious results greatly improved the quality of detection,avoided the blind detection of HBV DNA and reduced the experimental error that caused by reagent factors and non reagent factors.All the clinical samples for quantitative HBV DNA including the mutations of HBV gene can have been measured accurately and effectively,which was helpful to hepatitis B patients for antiviral therapy.