用于HIV-1 DNA检测的滤纸片干血斑能力验证质控品的制备及应用

Preparation and application of proficiency testing materials for HIV-1 DNA testing on dried blood spots

目的制备用于艾滋病病毒1型(HIV-1)DNA检测的干血斑能力验证质控品,并对其均一性和稳定性进行评估。方法培养仅含单拷贝的缺陷型HIV-1基因的8E5细胞,使用HIV-1阴性全血进行倍比稀释后,滴加到滤纸片上,制备成含有不同拷贝数8E5细胞的滤纸片干血斑。在不同温度条件下放置不同时间,然后使用雅培Real-time HIV-1定性检测试剂盒对其进行检测、整理并分析检测结果,评价滤纸片干血斑能力验证质控品的均一性和稳定性。结果滤纸片干血斑随机抽样平行检测结果完全一致;-20℃或4℃放置1年,室温放置3个月或在37℃放置4天,检测结果均一致。在应用性评价中,参加能力验证项目的15家实验室检测结果全部正确,制备的滤纸片干血斑作为质控品效果良好。结论使用8E5细胞制备的HIV-1DNA定性检测能力验证质控品的均一性、稳定性良好,是一种可用于目前中国HIV-1DNA定性检测能力验证质控品制备方法。

Objective To prepare proficiency testing materials of dried blood spots for HIV-1DNA proficiency testing,so as to guarantee HIV-1DNA test.Methods HIV-negative whole blood was spiked with a known number of freshly grown 8E5 cells,which contained a single integrated copy of HIV-1proviral DNA（defective genome）per cell.Positive samples and HIV-negative whole blood as negative samples were prepared by using Abbott RealTime HIV-1kit under different temperature conditions of preservation of filter paper with dried blood spots at different points of time for testing.Then the product stability and homogeneity of the DBS proficiency testing were evaluated.Results The sensitivity of the assay did not decrease after storage of the DBS for 1year at-20℃ or 4℃,for 3months at room temperature or for 4days after incubation at 37℃.All the Results of 15 laboratory participating in proficiency testing program were consistent with the expectations.Filter paper with dried blood spots prepared as proficiency testing materials work well.Conclusion The study shows that the stability and homogeneity of HIV-1DNA prepared by using 8E5 cells are satisfactory and it is an economic and convenient method to prepare the material for HIV-1DNA test.