建立梯度模型探讨溶血对TMA核酸检测的影响

Establishing a gradient model to explore the effect of hemolysis on TMA nucleic acid detection

目的通过建立溶血程度和病毒浓度的梯度模型,研究标本溶血对基于转录介导扩增(TMA)核酸检测(NAT)结果的影响。方法采用超声波破坏红细胞制造溶血标本,验证游离血红蛋白(Hb)浓度并建立溶血梯度模型;按照多因素实验设计中的析因设计方式,在不同溶血程度的标本模型中加入不同浓度的检测目标病毒。溶血程度根据血浆Hb浓度分为5个梯度,依次为0g/L、5g/L、10g/L、20g/L、40g/L。HIV、HCV、HBV浓度分为2个水平,低水平组分别为:20IU/ml、10IU/ml、5IU/ml;高水平组分别为:60IU/ml、20IU/ml、15IU/ml。采用基于TMA技术的全自动系统进行NAT。对检测数据进行收集、整理和统计学分析。结果本研究所建立的每个Hb梯度内各标本管的Hb浓度之间比较差异无统计学意义(P>α)。核酸检测分3个批次进行测试,每个批次80例样本,共计240例。其中225个测试标本呈反应性,15个对照标本呈非反应性。结论在标本血浆Hb浓度≤40g/L时,溶血对基于TMA技术的NAT结果无显著性影响。

Objective To investigate the effect of specimen hemolysis on NAT results based on TMA by establishing hemolysis level and virus concentration gradient model. Methods Ultrasound was used to destroy red blood cells in specimens,free Hb concentration was detected and hemolysis gradient model was established. According to the factorial design method in multi-factor experimental design,different concentrations of detection target viruses were added to the specimen models with different hemolysis levels. The degree of hemolysis was divided into 5 gradients according to plasma Hb concentration,which were 0 g/L,5 g/L,10 g/L,20 g/L,and 40 g/L,respectively. The concentrations of HIV,HCV and HBV were divided into two levels:low level group:20 IU/ml,10 IU/ml,5 IU/ml;high level group:60 IU/ml,20 IU/ml,15 IU/ml. NAT was implemented by an automatic system based on TMA technology. The test data was collected,organized and statistically analyzed. Results There was no statistically significant difference between the Hb concentrations of each specimen tube in each Hb gradient established in this study(P>α). Nucleic acid testing was conducted in three batches of 80 samples,a total of 240 samples. Of these,225 were reactive and 15 were non-reactive.Conclusion When the plasma Hb concentration of the specimen was ≤40 g/L,hemolysis had no significant effect on the TMAbased NAT results.