Standardization of Quantitative PCR for Human T-Cell Leukemia Virus Type 1 in Japan: a Collaborative Study
Standardization of Quantitative PCR for Human T-Cell Leukemia Virus Type 1 in Japan: a Collaborative Study
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DOI：
10.1128/jcm.01628-15
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发表时间：
2015-11-01
影响因子：
9.4
通讯作者：
Hamaguchi, Isao
中科院分区：
文献类型：
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作者：
Kuramitsu, Madoka;Okuma, Kazu;Hamaguchi, Isao
使用人类 T 细胞白血病病毒 1 型 (HTLV-1) 的定量 PCR (qPCR) 分析来评估整合到宿主血细胞基因组 DNA 中的 HTLV-1 原病毒 DNA 的量。越来越多的证据表明，高前病毒载量是发生成人 T 细胞白血病/淋巴瘤和 HTLV-1 相关脊髓病/热带痉挛性截瘫的危险因素之一。然而，qPCR 结果的实验室间差异使得评估实验室之间报告的原病毒载量的差异变得困难。为了纠正这种情况，我们试图通过合作研究中的 HTLV-1 qPCR 标准化来尽量减少实验室之间的差异。将携带 HTLV-1 原病毒的 TL-Om1 细胞用外周血单核细胞进行连续稀释，以制备候选标准品。通过统计评估标准品的原病毒载量以及每个实验室使用内部 qPCR 方法确定的原病毒载量，我们确定了实验室测量值与 TL-Om1 标准品理论值的相对比率。实验室的相对比率范围为0.84至4.45。接下来，我们使用相对比率校正 HTLV-1 携带者临床样本的前病毒载量。正如预期的那样，在应用校正后，实验室之间的总体差异减少了一半，从平均 7.4 倍减少到 3.8 倍。 HTLV-1 qPCR 可以使用 TL-Om1 细胞作为标准，并通过确定每个实验室中测量值与理论标准值的相对比率来标准化。
Quantitative PCR (qPCR) analysis of human T-cell leukemia virus type 1 (HTLV-1) was used to assess the amount of HTLV-1 provirus DNA integrated into the genomic DNA of host blood cells. Accumulating evidence indicates that a high proviral load is one of the risk factors for the development of adult T-cell leukemia/lymphoma and HTLV-1-associated myelopathy/tropical spastic paraparesis. However, interlaboratory variability in qPCR results makes it difficult to assess the differences in reported proviral loads between laboratories. To remedy this situation, we attempted to minimize discrepancies between laboratories through standardization of HTLV-1 qPCR in a collaborative study. TL-Om1 cells that harbor the HTLV-1 provirus were serially diluted with peripheral blood mononuclear cells to prepare a candidate standard. By statistically evaluating the proviral loads of the standard and those determined using in-house qPCR methods at each laboratory, we determined the relative ratios of the measured values in the laboratories to the theoretical values of the TL-Om1 standard. The relative ratios of the laboratories ranged from 0.84 to 4.45. Next, we corrected the proviral loads of the clinical samples from HTLV-1 carriers using the relative ratio. As expected, the overall differences between the laboratories were reduced by half, from 7.4-fold to 3.8-fold on average, after applying the correction. HTLV-1 qPCR can be standardized using TL-Om1 cells as a standard and by determining the relative ratio of the measured to the theoretical standard values in each laboratory.