Aubin J, Davi F, Nguyen-Salomon F, Leboeuf D, Debert C, Taher M, Valensi F, Canioni D, Brousse N, Varet B
Department of Hematology, CHU Necker Enfants-Malades, Paris, France.
Leukemia. 1995 Mar;9(3):471-9.
PCR amplification of IgH gene V-D-J junctional variability (IgH PCR) is increasingly replacing Southern analysis for the detection of clonal lymphoid populations in cases presenting diagnostic difficulties. In order to determine the most efficient strategy, we have compared three known methods, using consensus primers against the VH FR3 or FR2 (FR256) regions, or a mix of six primers against the FR1 region (FR1f), with a new approach using a consensus primer against FR1 (FR1c), never previously described for diagnostic purposes, on DNA from 89 monoclonal B-cell proliferations (16 ALL, 28 CLL/PLL, 15 myelomas, 30 NHL). We obtained a detection rate of 70% for FR3, 64% for FR1f and 77% and 78% for FR256 and FR1c, respectively. Polyclonal lymphocytes and mature T cell malignancies tested negative for all systems. Differences in the detection rate were related not only to the choice of VH primer but also the JH primer(s) used and the pathological subtype. All strategies led to adequate detection of leukaemic DNA, whereas the detection rate in myeloma varied between strategies from 47 to 80% and that of follicular lymphoma from 13 to 63%. The lowest detection rates were observed in follicular lymphoma and in mature CD5 negative proliferations, reflecting the probable correlation between somatic mutation and PCR false-negativity. The combined use of FR1c and FR256 allowed detection in at least one system of 92% of cases overall and at least 75% in all pathological subtypes, thus providing a simple, reliable and rapid non-radioactive system for the detection of B cell clonality.
免疫球蛋白重链（IgH）基因V-D-J连接区变异性的聚合酶链反应（PCR）扩增（IgH PCR）在诊断困难的病例中检测克隆性淋巴细胞群体方面，正越来越多地取代Southern印迹分析。为了确定最有效的策略，我们比较了三种已知方法，即使用针对VH FR3或FR2（FR256）区域的共有引物，或针对FR1区域的六种引物混合物（FR1f），以及一种使用针对FR1的共有引物（FR1c）的新方法（此前从未用于诊断目的），对来自89例单克隆B细胞增殖病例（16例急性淋巴细胞白血病、28例慢性淋巴细胞白血病/幼淋巴细胞白血病、15例骨髓瘤、30例非霍奇金淋巴瘤）的DNA进行检测。我们分别获得FR3的检测率为70%、FR1f为64%、FR256和FR1c分别为77%和78%。多克隆淋巴细胞和成熟T细胞恶性肿瘤在所有检测系统中均为阴性。检测率的差异不仅与VH引物的选择有关，还与所使用的JH引物以及病理亚型有关。所有策略均能充分检测白血病DNA，而骨髓瘤的检测率在不同策略之间为47%至80%，滤泡性淋巴瘤为13%至63%。滤泡性淋巴瘤和成熟CD5阴性增殖中检测率最低，这反映了体细胞突变与PCR假阴性之间可能存在的相关性。联合使用FR1c和FR256可使总体上至少92%的病例在至少一个系统中检测到，所有病理亚型中至少75%的病例可检测到，从而提供了一种简单、可靠且快速的非放射性检测B细胞克隆性的系统。
