丙型肝炎病毒核酸检测的国家标准物质的研制

Establishment of the first national standard for nucleic acid amplification technology (NAT) assay for HCV RNA

目的研制国内用于丙型肝炎病毒核酸（HCV RNA）扩增检测的血清标准物质.方法用HCV RNA阴性血浆将阳性血浆稀释至含量约300 000拷贝数/ml,然后进行真空冷冻干燥.检测方法采用实时荧光定量聚合酶链反应方法和罗氏公司的PCR内标定量方法.制备标准品含量通过与国际标准（世界卫生组织属下的国家生物标准研究所标准物质编号：96/790）比对获得.结果该标准物质定值为：（1.29±0.24）×10^5 IU/ml.其稳定性实验表明室温20～25℃（相对湿度20%～50%）14 d、37℃（相对湿度60%～80%）7 d均稳定.长期监测结果表明： 2～8℃6个月及-20℃保存两年的含量均无明显变化.均一性检测结果：瓶间不精密度为3.53%.结论该制备物可以作为用于核酸扩增检测的丙型肝炎病毒核酸国家标准物质.

Objective To establish an international standard for nucleic acid amplification technology（NAT） assay for HCV RNA in China. Methods Real-time PCR the candidate sample were HCV RNA positive and diluent with HCV-negative human plasma. The sample were then lyophilised with a concentration of approximately 300 000 copies/ml. The assay method used included the Roche Amplicor assay （version 2. 0 ） and real-time PCR. The lyophilised preparation were calibrated by the international standard （ NIBSC code ：96/790） from NIBSC. Results The titer of this lyophilised preparation is （ 1.29 ± 0. 24） × 10^5 IU/ml compare with the International Standard for HCV RNA 96/790. The stability test indicate that the sample were stable at room temperature（20-25℃ ） for 2 weeks and at 37℃ for at least 1 week. Long-term stability was abserved at 2-8℃ for 6 months and at -20℃ for more tan 2 years with no significant decrease. The vail-to-vail imprecision is 3.53%. Conclusions Based on the results of this study, the lyophilized sample was established as the first international standard for nucleic acid amplification technology （NAT） assay for HCV RNA in China.