Assay Design Affects the Interpretation of T-Cell Receptor Gamma Gene Rearrangements Comparison of the Performance of a One-Tube Assay with the BIOMED-2-Based TCRG Gene Clonality Assay
Assay Design Affects the Interpretation of T-Cell Receptor Gamma Gene Rearrangements Comparison of the Performance of a One-Tube Assay with the BIOMED-2-Based TCRG Gene Clonality Assay
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DOI：
10.2353/jmoldx.2010.090183
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发表时间：
2010-11-01
影响因子：
4.1
通讯作者：
Greiner, Timothy C.
中科院分区：
文献类型：
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作者：
Cushman-Vokoun, Allison M.;Connealy, Solomon;Greiner, Timothy C.
从多重荧光标记引物组获得的毛细管电泳结果的解释可能会因为小峰而变得复杂，这些小峰可能被错误地解释为克隆T细胞受体γ基因重排。在本报告中，使用了不同的分析设计来说明设计如何对特异性产生不利影响。DNA在所有三种连接引物上用相同的NED荧光染料扩增，首先结合（一种颜色试验），然后使用单个NED标记的连接引物单独扩增。单个引物试验设计表明，不显著的峰值很容易被错误地解释为克隆T细胞受体γ基因重排。两种方法的敏感性相似（92.9% vs 996%, P = 0.55），但特异性明显低于两种方法（88.5% vs 56.3%, P = 0.55）。使用5倍峰高比（P = 0.626），特异性提高到83%。这些发现表明，在解释具有多个独立分布的设计时需要格外谨慎，这比单一分布的分析更难解释（J Mol Diagn 2010, 12 787-796 DOI 10.2353/jmoldx.2010.090183）。
Interpretation of capillary electrophoresis results derived from multiplexed fluorochrome labeled primer sets can be complicated by small peaks, which may be incorrectly interpreted as clonal T cell receptor gamma gene rearrangements In this report, different assay designs were used to Illustrate how design may adversely affect specificity Ten clinical cases, with sub clonal peaks containing one of the two infrequently used joining genes, were identified with a tri color, one tube assay The DNA was amplified with the same NED fluorochrome on all three joining primers, first combined (one color assay) and then amplified separately using a single NED labeled joining primer The single primer assay design shows how insignificant peaks could easily be wrongly interpreted as clonal T cell receptor gamma gene rearrangements Next, the performance of the one tube assay was compared with the two tube BIOMED 2 based TCRG Gene Clonality Assay in a series of 44 cases Whereas sensitivity was similar between the two methods (92 9% vs 96 4%, P = 0 55), specificity was significantly less in the BIOMED 2 assay (87 5% vs 56 3%, P = 0 049) when a 2 x ratio was used to define clonality Specificity was improved to 81 3% by the use of a 5 x peak height ratio (P = 0 626) These findings illustrate how extra caution is needed in interpreting a design with multiple, separate distributions, which is more difficult to interpret than a single distribution assay (J Mol Diagn 2010, 12 787-796 DOI 10.2353/jmoldx.2010.090183)