摘要
目的 探讨不同样本处理方法对乳汁样本中HBV DNA定量结果的影响,筛选出适合临床实验室检测病毒DNA的样本前处理方法.方法 分别采用专用DNA提取液法、碱裂解法、酸化法、无水乙醇法和冷藏法,提取不同HBV DNA载量的乳汁样本中的HBV DNA,然后进行荧光定最PCR检测.结果乳汁样本中的HBV DNA〉10~4 IU/mL时,以专用DNA提取液法、碱裂解法、冷藏法处理的乳汁样本HBV DNA结果阳性率达100%,HBV DNA阳性血清用乳汁稀释与用阴性血清稀释的定量结果符合率100%,酸化法和无水乙醇法处理的乳汁样本HBV DNA均为阴性;当乳汁样本中的HBV DNA〈10~4 IU/mL时,以碱裂解法（NaOH浓度为0.4%～1.0%）处理的乳汁样本HBV DNA检测结果阳性率100%,HBV DNA阳性血清用乳汁稀释与用阴性血清稀释的定量结果符合率100%,其余方法处理的乳汁样本HBV DNA均为阴性.结论 碱裂解法提取乳汁样本中的HBV DNA适用临床样本检测,且NaOH浓度应控制在0.4%～1.0%之间.
Objective To investigate the pre-tretment influence of human breast milk specimens on the HBV DNA quantitative detection and optimize the methods of HBV DNA extraction from human breast milk specimens.Methods Different methods including DNA kit,Alkaline lysis,acidolysis,anhydrous ethanol and deepfreeze were used to extract HBV DNA from human breast milk specimens,with different HBV DNA load,and HBV DNA was quantitatively detected by real-time PCR.Results DNA kit,Alkaline lysis,deep freeze were equally efficient to extract HBV DNA,with the rate of positive at 100%,when the human breast milk contained HBV DNA of〉10~4 IU/mL.Moreover their coin-cidence rate of quantitative result was 100% in HBV DNA positive serum diluted by human breast milk and HBV DNA negative serum.While,other two methods led a negative result.However,only alkaline lysis（NaOH concentration was0.4%-1.0%）achieved a positive yield when the HBV DNA load of human breast milk specimens 〈10~4 IU/mL,and maintained the same coincidence rote of quantitative result at 100%.In contract,neither the other four pre-treatment methods gave a positive result.Conclunsions Alkaline lysis method can potentially adapted to clinical medicine for HBV DNA extraction from human breast milk specimens,with the NaOH concentration at 0.4%-1.0%.
出处
《国际流行病学传染病学杂志》
CAS
2010年第2期91-94,共4页
International Journal of Epidemiology and Infectious Disease
基金
杭州市科技局科技发展计划（20070733B02）
关键词
肝炎病毒
乙型
DNA
提取法
乳汁
Hepatitis B virus
DNA
Extraction
Human breast milk
