HBV-DNA定量检测核酸提取过程中影响因素的探讨  

Explore the influence factors of hepatitis B virus DNA quantitative testing in nucleic acid extraction process

作　　者：彭瑛[1] 温先勇[1] 邓正华[1] 

机构地区：[1]泸州医学院附属医院检验科,四川泸州646000

出　　处：《国际检验医学杂志》2015年第15期2156-2157,共2页International Journal of Laboratory Medicine

摘　　要：目的：探讨浓缩液剂量、沉淀混匀方式和裂解煮沸时间等前处理因素对实时荧光定量 PCR法检测乙型肝炎病毒DNA（HBV‐DNA）的影响。方法在浓缩液分别为90、95、105、110μL ；混匀方式分别为：漩涡混匀15 s组和30 s组，56℃预热120 s再漩涡混匀15 s组和56℃预热180 s再漩涡混匀15 s组；裂解煮沸时间分别为：6，8，12，14 min等条件下对80例自配的低值质控品，浓度在103～104 IU／mL进行测定，并与标准操作即浓缩液100μL ，镊子敲打混匀15 s ，裂解煮沸时间10 min比较。结果漩涡混匀15 s组和56℃预热180 s再漩涡混匀15 s与标准组比较差异有统计学意义（P＜0．05）；煮沸裂解时间6 min组与10 min组间比较差异有统计学意义（P＜0．05）。结论在一定范围内适当改变浓缩液剂量、混匀方式和煮沸裂解时间，不会影响检测结果。但经过一定时间预热处理后再振荡混匀的方式更容易使沉淀混匀和核酸的释放。Objective To investigate the concentrated liquid doses ,oscillation intensity and cracking boiling time to the influence of pre‐treatment HBV‐DNA .Methods The concentrate doses was 90 ,95 ,105 ,110 μL respectively ;mixing methods ：vortex mixing 15 seconds group and 30 seconds group ,56 ℃ preheated 120 seconds and then vortex mix 15 seconds group ,56 ℃ preheated 180 seconds and then swirl mix 15 seconds group ;cracking boiling time was 6 ,8 min ,under 12 min ,14 min respectively .Under the a‐bove conditions ,80 samples prepared by the laboratory were measured ,comparing with standard operating ,concentrate 100μL ,beat mix 15 seconds ,cracking boiling 10 min ,which HBV‐DNA concentration was 103 -104 IU/mL .Results Vortex mixing 15 s group and 56 ℃ preheat 180 s then swirl mixing 15 s were both a significant difference（P〈0 .05） in the mix mode groups ,comparing with the standard operating group .There were significant differences between the groups boiling lysis time of 6 min and 10 min group （ P〈0 .05） .Conclusion Within a certain range ,changing the dose of the concentrated solution ,mixing mode and the time of boiling lysis in extraction process of HBV‐DNA does not affect the test results ,But mixing ways after a certain period preheating make it easier to mix precipitation and release nucleic acid .

关 键 词：聚合酶链反应 浓缩液 振荡时间 煮沸时间 

分 类 号：R512.62[医药卫生—内科学]