全自动单核苷酸多态性分析仪检测JAK2基因V617F突变试验的性能评价  

Performance evaluation of the automated SNPs analyzer for the detection of JAK2 V617F mutation

作　　者：马玮哲 陈璐珈 康志华 许笑 陈宇明 张心菊 王俨 关明 Ma Weizhe;Chen Lujia;Kang Zhihua;Xu Xiao;Chen Yuming;Zhang Xinju;Wang Yan;Guan Ming(Central Laboratory, Huashan Hospital, Fudan University, Shanghai 200040, Chin)

机构地区：[1]复旦大学附属华山医院中心实验室,上海200040

出　　处：《中华检验医学杂志》2018年第6期470-474,共5页Chinese Journal of Laboratory Medicine

基　　金：上海中康医院发展中心市级医院临床辅助科室能力建设项目（SHDC22014001）

摘　　要：目的根据ISO15189认可要求评价全自动单核苷酸多态性分析仪i－densy IS－5320平台检测骨髓增殖性肿瘤患者JAK2基因V617F突变。方法仪器性能验证。选择2014年12月至2017年2月间复旦大学附属华山医院JAK2基因V617F突变阳性外周血标本20例，JAK2基因V617F突变阴性的外周血标本20例，以TaqMan MGB探针法实时荧光定量PCR作为对照方法，经测序方法验证评价仪器直接检测全血JAK2基因V617F突变的准确性。使用全血样本分别评估检测平台的检测重复性、携带污染与交叉污染，以及抗干扰能力。取HEL细胞与HCT116细胞为突变型对照和野生型对照，配制12种不同突变细胞比例样本进行检测，以此评价检测突变负荷的能力。结果i－densy IS－5320平台与TaqMan MGB探针实时荧光定量PCR方法检测完全一致；批内重复性与批间重复性为100%；未观察到携带污染与交叉污染；高胆红素（总胆红素：〉20.4 μmol/L）与高甘油三酯（甘油三酯：〉1.8 mmol/L）血样本对突变检测无明显干扰；该平台能够稳定检出负荷低至0.25%的突变。结论i－densy IS－5320可实现对全血直接进行JAK2基因V617F突变检测，具备较高的灵敏度、准确度和稳定性，操作简便，能够达到临床对突变的检测要求。Objective To test and evaluate the JAK2 gene V617F mutation in patients with myeloproliferative tumors based on i-densy IS-5320 platform according to ISO15189 accreditation requirements.Methods Instrument performance verification. Selected from December 2014 to February 2017, 20 cases of JAK2 V617F mutation positive peripheral blood samples from Huashan Hospital of the Shanghai FuDan University Medical College and 20 cases of peripheral blood samples with negative JAK2V617F mutation.The Realtime PCR with TaqMan MGB probe was selected as the control method to verify and evaluate the accuracy of testing JAK2 V617F mutation on i-densy IS-5320 platform. Whole blood samples were used to evaluate the reproducibility, cross-contamination and anti-interference ability of this platform. The ability of mutation load was verified by detecting mixtures of human erythnoleukemia cells and colorectal cancer cell HCT116 with 12 different proportions.Results I-densy IS-5320 platform and TaqMan MGB probe real-time fluorescence quantitative PCR show the same result.The within-run reproducibility and between-run reproducibility are both 100%. There is no observed contamination. High bilirubin and high triglyceride blood samples have no obvious interference on mutation detection. The mutation ratio with a load as low as 0.25% could be tested stably by i-densy IS-5320 platform. The detecting peak of melting curve can reflect the ratio of JAK2 V617F mutation to some extent.Conclusion sI-densy IS-5320 can detect the mutation of JAK2 V617F gene in the whole blood directly. It has high sensitivity, accuracy and stability, and is easy to operate, and also can reflect the mutation load of JAK2 V617F, which could meet the clinical requirements for the detection of mutations.

关 键 词：骨髓增殖性肿瘤 JAK2 V617F 高分辨率熔解曲线分析 

分 类 号：R733.3[医药卫生—肿瘤]