荧光定量PCR测定HBV DNA的线性评价和临床可报告范围  

Linearity evaluation and clinical reportable range of HBV DNA determination by fluorescence quantitative PCR

作　　者：李育敏 阚丽娟 张水兰[1] 汤花梅 熊丹 张秀明 LI Yumin;KAN Lijuan;ZHANG Shuilan;TANG Huamei;XIONG Dan;ZHANG Xiuming(Department of Clinical Laboratory,People′s Hospital of Luohu District,Shenzhen,Guangdong 518001,China)

机构地区：[1]广东省深圳市罗湖区人民医院医学检验科

出　　处：《检验医学与临床》2019年第20期2952-2955,共4页Laboratory Medicine and Clinic

基　　金：广东省深圳市医疗卫生三名工程（SZSM201601062）

摘　　要：目的探讨荧光定量PCR测定乙型肝炎病毒核酸(HBV DNA)检测系统的线性评价和临床可报告范围(CRR)。方法选取接近线性范围上限的患者标本(8.81E+08)IU/mL,用阴性标本进行系列稀释至比厂商声明的线性范围下限低一个数量级(8.81E+01)IU/mL,共制备8种浓度梯度的标本,每个标本重复测量3次,在一个分析批内完成,采用平均斜率法、美国临床和实验室标准化委员会EP 06-A指南方法和线性稀释回收法进行线性评价。在线性范围内选取3份高浓度患者标本,采用最大稀释度试验确定CRR。结果采用平均斜率法,相关系数(r)=0.999,斜率(b)=1.004(t=93.326,P=0.000),符合线性要求;按EP 06-A指南,经多项式回归分析,最适多项式为二次,二次多项式中每种浓度的线性偏差均小于允许误差0.4(log值);采用线性稀释回收法,r=0.999,b=0.993(t=75.82,P=0.000),符合线性要求,各浓度梯度的稀释回收量均≤±0.4(log值)。采用最大稀释度试验确定最大稀释倍数为1∶100 000。结论HBV DNA在(8.81E+01)^(8.81E+08)IU/mL为临床可接受线性关系,CRR为(8.81E+01)^(8.81E+13)IU/mL。Objective To explore the linearity evaluation and clinical reportable range(CRR)in HBV DNA determination by fluorescence quantitative PCR.Methods A patient′specimen with an analytic concentration at(8.81E+08)IU/mL which was near the expected upper linear range limit was diluted to a concentration at the expected lower limit(8.81E+01)IU/mL with a negative specimen.A series of samples with 8 equally spaced concentrations were provided and each sample was tested 3 times at each level within one run.Average slope method,the method recommended by CLSI EP 06-A and linear dilution recovery method were used for the linearity evaluation.Three higher specimens were selected for CRR determination by the maximum valid dilution method.Results The results analyzed by average slope method indicated that the correlation coefficient(r)was 0.999,the slope(b)was 1.004(t=93.326,P=0.000),and which met the linear standard.The results analyzed by EP 06-A indicated that second-order model had the best fit in the models by polynomial regression analysis.None of the deviation from linearity at each concentration in second-order model exceeded the allowable error(log 0.4).The results analyzed by linear dilution recovery method indicated that r was 0.999,b was 0.993(t=75.82,P=0.000),and which met the linear standard.The amount of recovery at each level was≤±log 0.4.The results analyzed by the maximum valid dilution method indicated that the maximum dilution ration was 1∶100 000.Conclusion The HBV DNA concentration which ranges from(8.81 E+01)IU/mL to(8.81 E+08)IU/mL is a accepted linear range in clinic.CRR ranges from(8.81E+01)IU/mL to(8.81E+13)IU/mL.

关 键 词：乙型肝炎病毒核酸 线性评价 临床可报告范围 平均斜率法 

分 类 号：R446.1[医药卫生—诊断学]