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人乳腺癌特异性基因1 mRNA实时荧光定量逆转录-聚合酶链反应检测试剂盒的建立及应用
引用本文:程民,沈国栋,卞庚,徐婷娟,徐维平,沈干,胡世莲.人乳腺癌特异性基因1 mRNA实时荧光定量逆转录-聚合酶链反应检测试剂盒的建立及应用[J].中国临床保健杂志,2014(1):63-67,I0003.
作者姓名:程民  沈国栋  卞庚  徐婷娟  徐维平  沈干  胡世莲
作者单位:[1]安徽医科大学附属省立医院、安徽省立医院,合肥230001 [2]安徽省老年医学研究所 ,合肥230001 [3]肿瘤免疫与营养治疗安徽省重点实验室,合肥230001
基金项目:国家自然科学基金面上项目(81071808);安徽省国际科技合作项目(10080703037);安徽省科技攻关项目(12010402126);安徽省肿瘤免疫营养诊疗产业创新团队资助
摘    要:目的建立人乳腺癌特异性基因1(BCSG1)mRNA实时荧光定量逆转录-聚合酶链反应(RTPCR)检测试剂盒,并评价其在乳腺癌循环肿瘤细胞检测中的应用价值。方法优化试剂盒中各组分的浓度、确定试剂盒的稳定性、灵敏度及特异性,并检测外周血样品中BCSG1 mRNA的表达水平。结果优化了试剂盒中的引物、探针,成功制备了定值标准品,通过设立阴、阳性质控品的方法,解决了试剂盒的特异性、灵敏性、准确性及质量控制等关键技术问题,特异度及准确性均为95%以上,灵敏度为100拷贝/ml(全血)。外周血标本中BCSG1 mRNA的检测结果为,12例临床确诊已转移的乳腺癌患者均为阳性,21例临床未确诊转移的乳腺癌患者中12例为阳性,其余9例为阴性,15例乳腺良性疾病患者、10例正常人均为阴性。结论建立的BCSG1 mRNA实时荧光定量RT-PCR检测试剂盒具有很高的临床应用价值,可用于检测外周血样品中BCSG1mRNA的表达量,检测结果可作为乳腺癌患者诊疗过程中的重要监测指标。

关 键 词:乳腺肿瘤  试剂盒  诊断  RNA  信使  逆转录聚合酶链反应

Establishment and application of a detection kit for human breast cancer-specific gene 1 mRNA by real-time ;quantitative RT-PCR
CHEN Min,SHEN Guodong,BIAN Geng,XU Tingjuan,XU Weiping,SHEN Gan,HU Shilian.Establishment and application of a detection kit for human breast cancer-specific gene 1 mRNA by real-time ;quantitative RT-PCR[J].Chinese JOurnal of Clinical Healthcare,2014(1):63-67,I0003.
Authors:CHEN Min  SHEN Guodong  BIAN Geng  XU Tingjuan  XU Weiping  SHEN Gan  HU Shilian
Institution:(Anhui Province Hospital Affiliated Anhui Medical University ,Hefei 230001, China)
Abstract:Objective This study was to establish a quantitative method for human breast cancer-specific gene 1(Breast cancer-specific Gene 1,BCSG1)mRNA by real-time RT-PCR.Its application was further evaluated by detecting circulation tumor cells in the peripheral blood of breast cancer patients .Method The concentration of each component in the detection kit was optimized;the stability,sensitivity and specificity were determined .The expression level of BCSG1 mRNA was detected in the samples of peripheral blood .Results Optimized primers and probes were obtained,and setting standards were successfully prepared .Through establishing the positive and negative control ma-terials,the specificity,sensitivity,accuracy and quality control were determined for the detection kit .The specificity and accuracy were both more than 95%,and the sensitivity was of 100 copies/ml( whole peripheral blood ) .The detec-tion results of BCSG1 mRNA in peripheral blood samples showed that all the samples were positive for 12 breast canc-er patients clinically diagnosed with metastasis;and 12 samples were positive and other 9 samples were negative among 21 breast cancer patients without clinically diagnosed with metastasis or not ;all the samples were negative for 15 pa-tients with benign breast disease;and all the samples were negative for 10 healthy control .Conclusion The quantita-tive RT-PCR assay for BCSG1 mRNA could be used for the detection of circulating tumor cells in peripheral blood of breast cancer patients ,which would be valuable for the diagnosis and clinical-decision making for breast cancer patients .
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