3种RNA提取方法在血清丙型肝炎病毒RNA定量检测中的性能评价  

Three Commercial RNA Extraction Methods in Hepatitis C Viral Load Test:Their Comparative Evaluation

作　　者：张磊[1] 陆小军[1] 应斌武[1] 庄杰[1] 丁柳[1] 代波[1] 王军[1] 刘显中[1] 林立[1] 宋兴勃[1] 

机构地区：[1]四川大学华西医院实验医学科分子诊断室,四川成都610041

出　　处：《中华医院感染学杂志》2009年第22期3027-3029,共3页Chinese Journal of Nosocomiology

摘　　要：目的比较3种方法对血清HCV RNA的提取性能,并对其作方法学评价,揭示RNA提取技术对HCVRNA定量的影响。方法使用QIAamp Viral RNA Mini Kit(Q法),NucliSens miniMag系统(M法)和匹基(PG)丙型肝炎病毒定量试剂盒自带RNA提取方法(P法)进行血清HCV RNA提取,实时荧光RT PCR定量检测,对77例临床血清标本进行了提取检测,同时设计了方法学评价实验,从准确度,重复性,线性和灵敏度方面评价3种方法。结果77例临床标本的M、Q和P法阳性率分别是85.7%、84.4%和36.3%;25例共同阳性标本中,M和Q法的CT值分别比P法平均低7.5和9.35,表现出高于P法的提取效率和RNA质量;M和Q法的标准变异指数(S.D.I.)绝对值均<2,P法S.D.I.绝对值>2;M法平均CV为40.0%,Q法为71.0%,P法>100.0%;M法表现最佳的线性(R>0.99),M、Q和P法的最低检测限分别为39.4、394和3943 IU/ml。结论从提取效率和收获RNA质量上M和Q法远远优于P法,进一步方法性能表明,M法为HCV RNA定量最佳提取方法,而P法因其提取效率低、收获RNA质量差以及重复性不好,不适合临床HCV RNA定量试验。OBJECTIVE To perform a comparative validation on three systems：QIAamp Viral RNA Mini Kit （Q）, the NucliSens miniMAG extraction system （M）, the PG method （P）, in an attempt to elucidate the impact of extraction methods on the quantitative assay of HCV RNA in serum. METHODS A total of 77 anti-HCV positive serum specimens submitted for hepatitis viral load determination were used. Method performance： accuracy, reproducibility, linearity and sensitivity were evaluated. RESULTS In 77 serum specimens, the HCV RNA was detected at 85. 7%, 84. 4% and 36. 3% in nucleic acid extracts prepared by the methods M, Q and P, respectively. The M and Q produced lower CT values in real time Lightcycler PCR than P （average difference was -7.5 and -9. 35 ）, thus showed the higher efficiency of extraction and quantity of RNA compared to P. The standard deviation index （S. D. I. ） was less than 2 for M and Q and more than 2 for P. The method M showed the best precision and linearity among the three systems （mean CV, 40%; R〉0.99）. The lowest detection limit was 39.4, 394, and 3940 IU/ml for M, Q and P, respectively. CONCLUSIONS The M and Q exhibit much higher extraction efficiency and quality of RNA than P. With further evidence from method evaluation M is considered to be the best for HCV RNA quantitative assay. It also suggest that the P be not suitable to HCV RNA quantitative assay due to the low extraction efficiency, poor quality of yielding RNA and reproducibility.

关 键 词：RNA提取 HCV RNA定量 提取效率 RNA质量 CT值 

分 类 号：R446[医药卫生—诊断学]