智能等温扩增方法建立乙肝病毒核酸检测体系及评估

Establishment and evaluation of the smart Amplification Process for Hepatitis B Virus DNA

目的:利用智能等温扩增方法(SMAP)建立乙型肝炎病毒(HBV)核酸检测体系。方法:以pGEMX-T-X重组质粒为标准品,应用SMAP、实时荧光定量PCR法(Real-time PCR)对标准品的连续梯度稀释样品和其他几种病毒核酸提取物进行对照试验,测定SMAP对HBV核酸检测的特异性与灵敏度。结果:HBV病毒SMAP检测体系,反应最短时间15min,最优反应时间30min,无特殊仪器要求,特异性好,灵敏度10copies/μl;HBV病毒Real-time PCR检测体系,反应时间约2h,需要实时荧光定量PCR仪,特异性好,灵敏度100copies/μl。结论:SMAP方法检测速度快,无特殊仪器要求,能够提高检查效率,降低诊疗成本,值得推广应用。

Objective： Used SMAP （the Smart Amplification Process）to establish HBV-DNA detection methods. Methods： Used pGEMXTX plasmid as standard sample to establish HBV-DNA detection methods （including Real-time PCR and SMAP） and set a series of control trials to compare their sensitivity and specificity. Results： The shortest reaction time and optimal reaction time of SMAP method are 15min and 30rain separately. This method needs no special instrument, but a stable reaction temperature of 65℃. There was no cross react with other viruses. The sensitivity of SMAP method was 10copies/μl; the reaction of Real-time PCR method costs 2 hours and needs special instrument. There was no cross react with other viruses. The sensitivity of real-time PCR method was 100copies/μl. Conclusion： SMAP method for HBV-DNA detecting had an advantage of fast detected speed and no requirement for special instrument. It will help to increase the detecting effect and decrease the detecting cost. So it worth to be popularized and applied in clinic.