Bootman J S, Kitchin P A
MRC PCR Reference Centre, AIDS Collaborating Centre, NIBSC, Potters Bar, Herts, UK.
J Virol Methods. 1992 Apr;37(1):23-41. doi: 10.1016/0166-0934(92)90018-9.
An international collaborative study was performed to evaluate a set of PCR reference reagents for HIV diagnosis. Twenty-six laboratories from 9 countries analysed a proficiency panel of 10 coded DNA samples using the PCR reference reagents and protocols. For comparison, these coded samples were then assessed using a laboratory's own 'in-house' reagents and methodologies. The objectives of the study were: (i) to assess inter-laboratory variation of PCR sensitivity, (ii) to evaluate the DNA 'carryover' problem and frequency of false negative results and (iii) to examine the utility of the complete set of reagents and templates to act as reference preparations for HIV PCR. Using the reference reagents, 46% of laboratories reported no false positive results in any of their assays of the negative controls. The remaining laboratories all reported a false positive result(s) in at least one assay. The overall false positive result rate for the study was 9.3%. In contrast, an overall false negative result rate of 7.4% was observed, with some laboratories recording negative results even for samples containing 10,000 molecules of target DNA. The level of absolute sensitivity may be assessed accurately only from the 12 laboratories that obtained no false positive results. All 12 laboratories detected the sample containing 10 molecules of template DNA and 9 out of the 12 laboratories detected the sample containing 1 molecule. This is in close agreement with the theoretical detection rate based on a statistical probability model for the detection of a single molecule. These characterised reference reagents were at least as sensitive as any of the 'in-house' reagents and methodologies applied, including nested PCR. The complete set of characterised reference reagents is now available for quality control assessment of HIV-1 PCR from the MRC ADP.
开展了一项国际合作研究，以评估一组用于HIV诊断的PCR参考试剂。来自9个国家的26个实验室使用这些PCR参考试剂和方案分析了一个包含10个编码DNA样本的能力验证样本组。为作比较，随后使用各实验室自己的“内部”试剂和方法对这些编码样本进行评估。该研究的目的是：(i)评估PCR敏感性的实验室间差异；(ii)评估DNA“残留”问题以及假阴性结果的频率；(iii)检验整套试剂和模板作为HIV PCR参考制剂的效用。使用参考试剂时，46%的实验室报告称其对阴性对照的任何检测中均未出现假阳性结果。其余实验室均报告至少在一次检测中出现了假阳性结果。该研究的总体假阳性结果率为9.3%。相比之下，观察到总体假阴性结果率为7.4%，一些实验室甚至对含有10,000个目标DNA分子的样本检测出阴性结果。只有从12个未出现假阳性结果的实验室才能准确评估绝对敏感性水平。所有12个实验室都检测到了含有10个模板DNA分子的样本，12个实验室中有9个检测到了含有1个分子的样本。这与基于单分子检测统计概率模型的理论检测率密切相符。这些经过特性鉴定的参考试剂至少与所应用的任何“内部”试剂和方法（包括巢式PCR）一样敏感。现在可从MRC ADP获得整套经过特性鉴定的参考试剂，用于HIV-1 PCR的质量控制评估。
