冷冻保存血清对HBV-DNA荧光定量PCR检测结果的影响

Impact of cryopreservation of serum on test result of HBV-DNA by fluorescence quantitative PCR

目的 研究荧光定量PCR检测一年内冷冻保存（-80℃）血清中HBV-DNA的结果 与冷冻保存时间的相关性。方法 采集25例乙型肝炎患者血清标本,将其分成4份于-80℃保存,分别在第0、2、6、12月相同条件下冻融,再进行HBV-DNA荧光定量PCR检测。将检测数据转化成对数值,采用分析软件处理数据。结果 25份血清标本HBV-DNA初始值为103-107copies/ml,2个月冷冻保存的血清各组标本HBV-DNA含量较初始值没有明显差异（P=0.39）;但在冷冻保存6个月和12个月后HBV-DNA含量显著低于初始值（P=0.01,P=0.00）;6个月冷冻保存也明显低于2个月保存HBV-DNA含量（P=0.014）。一元线性回归分析显示,HBV-NDA检测的含量与时间成负相关（r=0.985）。结论 冷冻保存（-80℃）血清时间越长HBV-DNA含量随之越低。

Objective To study the impact of cryopreservation（ - 80 ℃ ） of serum on test result of HBV-DNA by fluores- cence quantitative PCR and the relationship between the DNA degradation and the cryopreservation time. Methods The serum samples of 25 patients with hepatitis B were collected and divided into four tubes to be stored at - 80 ℃. The fluorescence quantitative PCR was performed at 0,2,6,12 months after the cryopreservation to detect HBV-DNA（ the condi- tions of freezing and thawing were same）. The testing Data is converted to logarithm values. ANOVA and linear regression analysis were performed. Results The initial value of HBV-DNA in 25 specimens were 103 - 107 copies/ml,and did not decreased obviously in 2 months（P = 0.39） ;The cryopreservation of 6 and 12 months significantly decreased the content of HBV-DNA as compared with the initial value（ P = 0.01 ,P = 0. 00） and the values at 2 months（ P = 0. 014）. Linear regression analysis showed a negative correlation between the cryopreservation time and the content of HBV-NDA （r = 0.985 ）. Conclusion The long time（ one year or more） cryopreservation of serum can promote the HBV-DNA degradation and has a bad influence on the test results.