中文摘要：

目的：初步探讨荧光定量PCR溶解曲线分析技术监测人外周血T细胞TCRalpha链CDR3谱系漂移（单/寡/多克隆增生）。方法：提取4例正常人、2例淋巴瘤型白血病患者PBMC中的总RNA,逆转录成cDNA,以32个人TCRalpha链胚系可变区基因家族（TRAV）设计上游引物,共同的TCRalpha胚系链恒定区基因家族（TRAC）设计下游引物,荧光定量PCR（FQ-PCR）扩增32个TRAV基因各家族CDR3谱系,溶解曲线法分析各家族CDR3谱系的单/寡/多克隆增生。结果：正常人外周血T细胞TCRalpha链32个家族CDR3表达频率不一致,各家族PCR产物的溶解曲线谱型图（melting curve spectratyping）呈现熔点不同的CDR3多态性,为多克隆增生的高斯分布,2例淋巴瘤型白血病患者外周血T细胞TCRalpha链32个家族CDR3表达频率不一致,部分家族呈缺失状态,患者各家族PCR产物的溶解曲线谱型图上,多数家族为多克隆增生的高斯分布,但每个患者均出现数量不等的单克隆和寡克隆增生家族。结论：荧光定量PCR溶解曲线分析TCRalpha链CDR3谱系漂移技术方法稳定简便,能较好地监测正常人和临床样本外周血T细胞TCRalpha链CDR3谱系漂移（单/寡/多克隆增生）。

英文摘要：

Objective：To establish the technique for real-time fluorescence quantitative reverse transcription polymerase chain reaction （FQ-PCR） by DNA melting curve analysis for detecting the CDR3 shewing of TCR alpha gene repertoire in human peripheral blood. Methods： Total RNA of peripheral blood mononuclear cell （PBMC） from 4 healthy donors and 2 patients with lymphomatous leukemia were transcripted reversely into cDNA. The cDNA of 32 TRAV gene family CDR3 was amplified by FQ-PCR. Analysis of the monoclonal/oligeclonal/polyclonal CDR3 spectratyping with DNA melting curve. Results：The FQ-PCR products of 32 TRAV family CDR3 were showedas a blur land at the predicted of products size in healthy donors and parts of TRAV family CDR.3 products disappeared in patients on 1.5% agarose gel by Gold-View staining. The 32 TRAV family CDR3 were showed with different frequencies by relative fluorescence quantitative in healthy donors and the patients. The CDR3 spetratyping for 32 TRAV families was showed as polyclonal peak （ Gaussian distribution） in healthy donors but showed as different monoclonaL/ohgoclonal/polyclonal peak in the patients with lymphomatous leukemia with DNA melting curve analysis（ we called ＂melting curve spectratyping of CDR3＂）Conclusion：The study suggests that the technique of ＂FQ-PCR with DNA melting curve analysis be convenience and celerity for detecting the CDR3 skewing of TCR alpha gene repertoire in human peripheral blood.