实时定量PCR法测定乙肝病毒脱氧核糖核酸  

Real-time PCR Assay for Quantification of Hepatitis B Virus DNA

作　　者：王蔚青[1] 王彤[1] 孟玲[1] 杜青[1] 黄佩珺[1] 周思蒙 

机构地区：[1]南京医科大学第一附属医院,南京210029 [2]艾克斯马赛第三大学,法国13003

出　　处：《药学与临床研究》2010年第2期149-151,155,共4页Pharmaceutical and Clinical Research

摘　　要：目的:建立实时定量PCR法测定血清样品中乙肝病毒脱氧核糖核酸(HBV-DNA)含量。方法:用PEG-裂解-煮沸法从血清中提取核酸,制作成标准浓度曲线后,对100份患者样本进行实时荧光定量分析,并与ELISA(酶标法)结果作比较,再与CAP-CTM法(化学发光法)进行比较。结果:用SPSS(V.16)软件进行统计分析,得线性回归方程Y=-3.212X+44.90;r2=0.9980;日内、日间平均误差为RSD<2.15%;最低检测限是400copies/mL(拷贝数);该法测得患者HBV-DNA的浓度与ELISA测出样品阳性发生率呈正相关,再与CAP-CTM作配对t检验,P>0.05,两者无显著性差异。结论:新建的PCR检测法敏感度高,重现性好,线性范围较广,可用于进行临床血清样本中HBV-DNA的定量检测,这对评估HBV患者诊治效果有帮助。Objective： To establish a real-time PCR method to determine HBV DNA concentration in human serum. Methods： We utilized PEG-lyses-boiling method to extract HBV DNA from the serum sample, Seven standard samples were used to determine the linearity of the assay. The realtime PCR was applied on 100 cases of serum, two other methods were applied for comparison. Results： Statistical analysis was done with the SPSS statistical of software（versionl6）; A good linearity was seen in the calibration curves with r^2=0.9980. Linear equation of regression was Y=-3.212X＋ 44.90. The intra-day and inter-day precision （RSD） was〈2.15%. The minimum detection limit was 400 copies/mL. Quantitative assay of the real-time PCR has positive relevance to the HBV DNA positive rate of ELISA test, but show no significant difference from t.he CAP-CTM test （P〉0.05）. Conclusions： This method can be used for the determination of HBV DNA concentration in human serum and for therapeutic assessment in HBV patients.

关 键 词：乙肝病毒脱氧核糖核酸 实时荧光定量扩增法 酶标测定法 

分 类 号：R392-33[医药卫生—免疫学]