中华实验和临床病毒学杂志
中華實驗和臨床病毒學雜誌
중화실험화림상병독학잡지
CHINESE JOURNAL OF EXPERIMENTAL AND CLINICAL VIROLOGY
2012年
2期
142-144
,共3页
郑惠惠%邱丰%曾经瑗%毕胜利
鄭惠惠%邱豐%曾經瑗%畢勝利
정혜혜%구봉%증경원%필성리
肝炎病毒,甲型%逆转录聚合酶链反应%敏感性与特异性
肝炎病毒,甲型%逆轉錄聚閤酶鏈反應%敏感性與特異性
간염병독,갑형%역전록취합매련반응%민감성여특이성
Hepatitis A virus%Reverse traseriptase polymerace chain reaction%Sensitivity and specificity
目的 建立甲肝病毒(HAY)特异性TaqMan荧光定量PCR检测方法,并对血清等样品中的HAV进行检测.方法 根据参考文献,选取HAV基因组保守区5'-NCR区引物和探针,建立特异性强、敏感度高、稳定性好的TaqMan HAV Real-time RT-PCR检测体系并应用于甲肝患者血清等病毒核酸的检测.结果 本研究建立的方法能够特异检测出样品中的甲肝病毒,其灵敏度介于每反应0.1CCID50至0.01CCID50之间.同一样本重复检测3次,批内样本Ct值的变异系数最大2.0％,批间样本Ct值的变异系数最大2.6％.急性期甲型肝炎患者血清中甲肝病毒为5.18×102～4.93×107拷贝/ml.结论 本实验建立的HAV Real-time RT-PCR检测方法具有特异、灵敏、快速等优点,可成功应用于临床样本等甲肝病毒的病原学检测.
目的 建立甲肝病毒(HAY)特異性TaqMan熒光定量PCR檢測方法,併對血清等樣品中的HAV進行檢測.方法 根據參攷文獻,選取HAV基因組保守區5'-NCR區引物和探針,建立特異性彊、敏感度高、穩定性好的TaqMan HAV Real-time RT-PCR檢測體繫併應用于甲肝患者血清等病毒覈痠的檢測.結果 本研究建立的方法能夠特異檢測齣樣品中的甲肝病毒,其靈敏度介于每反應0.1CCID50至0.01CCID50之間.同一樣本重複檢測3次,批內樣本Ct值的變異繫數最大2.0％,批間樣本Ct值的變異繫數最大2.6％.急性期甲型肝炎患者血清中甲肝病毒為5.18×102～4.93×107拷貝/ml.結論 本實驗建立的HAV Real-time RT-PCR檢測方法具有特異、靈敏、快速等優點,可成功應用于臨床樣本等甲肝病毒的病原學檢測.
목적 건립갑간병독(HAY)특이성TaqMan형광정량PCR검측방법,병대혈청등양품중적HAV진행검측.방법 근거삼고문헌,선취HAV기인조보수구5'-NCR구인물화탐침,건립특이성강、민감도고、은정성호적TaqMan HAV Real-time RT-PCR검측체계병응용우갑간환자혈청등병독핵산적검측.결과 본연구건립적방법능구특이검측출양품중적갑간병독,기령민도개우매반응0.1CCID50지0.01CCID50지간.동일양본중복검측3차,비내양본Ct치적변이계수최대2.0％,비간양본Ct치적변이계수최대2.6％.급성기갑형간염환자혈청중갑간병독위5.18×102～4.93×107고패/ml.결론 본실험건립적HAV Real-time RT-PCR검측방법구유특이、령민、쾌속등우점,가성공응용우림상양본등갑간병독적병원학검측.
Objective To establish a specific TaqMan-based Real-time PCR assay for the detection of hepatitis A virus in serum samples.Methods According to the references,primers-probe sets which were located in 5'-NCR,the most conservative part of HAV genome were designed and therefore we established a TaqMan real-time RT-PCR assay with great performance of specificity, sensitivity and reproducibility.And then it was used in the detection of HAV RNA in serum from HAV patients.Results The HAV Real-time RT-PCR assay established in this study were able to detect HAV RNA and its detection limit ranged from 0.1CCID50/reaction to 0.01CCID50/reaction.When the detection of a same sample was repeated for three times,coefficients of varistion (CV) of intra- and inter-assay were calculated and they were all less than 2.0％ and 2.6％ respectively.Our data suggested that there were 5.18 × 102 - 4.93 × 107 RNA copies in 1 ml of the serum from acute HAV patients.Conclusion The TaqMan-based Real-time PCR assay established in this study was specific and precise for the rapid detection of HAV RNA.It was applied successfully in the pathogen detection of clinical samples.
