Jones Carol D, Yeung Cecilia, Zehnder James L
Department of Pathology, Stanford University School of Medicine, Stanford, CA, USA.
Am J Clin Pathol. 2003 Jul;120(1):42-8. doi: 10.1309/60A9-C8WG-EGHR-NXEE.
We developed and extensively validated a real-time PCR assay for the quantitation of bcr-abl to determine residual disease in patients with chronic myelogenous leukemia. This method quantitates the p210 and the p190 bcr-abl RNA fusion transcripts with results normalized to a housekeeping gene, using the 5'-exonuclease technique and the ABI PRISM 7700 Sequence Detection System (Applied Biosystems, Foster City, CA). We parallel tested 372 clinical specimens and 50 peripheral blood samples from patients not known to have any myeloproliferative disorders. The results were 100% specific. Sensitivity studies showed that this method can detect bcr-abl in cell lines diluted to 0.0001% and can detect a single bcr-abl plasmid spiked into negative RNA. The between-run reproducibility showed a coefficient of variance (CV) of 12.3%, and within-run reproducibility showed a CV of 13.8%. This method can be used to reliably monitor the disease load in patients with bcr-abl-positive diseases.
我们开发并广泛验证了一种用于定量检测bcr-abl的实时聚合酶链反应（PCR）检测方法，以确定慢性粒细胞白血病患者的残留疾病。该方法使用5'-外切核酸酶技术和ABI PRISM 7700序列检测系统（应用生物系统公司，加利福尼亚州福斯特城），对p210和p190 bcr-abl RNA融合转录本进行定量，结果以管家基因进行标准化。我们对372份临床标本和50份来自未知患有任何骨髓增殖性疾病患者的外周血样本进行了平行检测。结果具有100%的特异性。敏感性研究表明，该方法能够检测稀释至0.0001%的细胞系中的bcr-abl，并且能够检测掺入阴性RNA中的单个bcr-abl质粒。批间重复性显示变异系数（CV）为12.3%，批内重复性显示CV为13.8%。该方法可用于可靠地监测bcr-abl阳性疾病患者的疾病负荷。
