Hourfar M K, Schmidt M, Seifried E, Roth W K
Institute of Transfusion Medicine and Immunohaematology, German Red Cross, Johann Wolfgang Goethe University, Frankfurt, Germany.
Vox Sang. 2005 Aug;89(2):71-6. doi: 10.1111/j.1423-0410.2005.00649.x.
Nucleic acid extraction still harbours the potential for improvements in automation and sensitivity of nucleic acid amplification technology (NAT) testing. This study evaluates the feasibility of a novel automated high-volume extraction protocol for NAT minipool testing in a blood bank setting.
The chemagic Viral DNA/RNA Kit special for automated purification of viral nucleic acids from 9.6 ml of plasma by using the chemagic Magnetic Separation Module I was investigated. Analytical sensitivity for hepatitis C virus (HCV), human immunodeficiency virus-1 (HIV-1), hepatitis B virus (HBV), hepatitis A virus (HAV) and parvovirus B19 (B19) was compared to our present manual procedure that involves virus enrichment by centrifugation.
Chemagic technology allows automation of the viral DNA/RNA extraction process. Viral nucleic acids were bound directly to magnetic beads from 9.6-ml minipools. By combining the automated magnetic beads-based extraction technology with our in-house TaqMan polymerase chain reaction (PCR) assays, 95% detection limits were 280 IU/ml for HCV, 4955 IU/ml for HIV-1, 249 IU/ml for HBV, 462 IU/ml for HAV and 460 IU/ml for B19, calculated for an individual donation in a pool of 96 donors. The detection limits of our present method were 460 IU/ml for HCV, 879 IU/ml for HIV-1, 90 IU/ml for HBV, 203 IU/ml for HAV and 314 IU/ml for B19.
The 95% detection limits obtained by using the chemagic method were within the regulatory requirements for blood donor screening. The sensitivities detected for HCV, HBV, HAV and B19 were found to be in a range similar to that of the manual purification method. Sensitivity for HIV-1, however, was found to be inferior for the chemagic method in this study.
核酸提取在核酸扩增技术（NAT）检测的自动化和灵敏度方面仍有改进的潜力。本研究评估了一种新型自动化大容量提取方案在血库环境中用于NAT混合样本检测的可行性。
研究了chemagic病毒DNA/RNA试剂盒，该试剂盒专门用于通过chemagic磁分离模块I从9.6毫升血浆中自动纯化病毒核酸。将丙型肝炎病毒（HCV）、人类免疫缺陷病毒1型（HIV-1）、乙型肝炎病毒（HBV）、甲型肝炎病毒（HAV）和细小病毒B19（B19）的分析灵敏度与我们目前采用离心富集病毒的手动方法进行了比较。
chemagic技术可实现病毒DNA/RNA提取过程的自动化。病毒核酸直接从9.6毫升混合样本中与磁珠结合。通过将基于磁珠的自动化提取技术与我们内部的TaqMan聚合酶链反应（PCR）检测相结合，对于96名献血者组成的混合样本中的单个献血者，计算得出HCV的95%检测限为280 IU/ml，HIV-1为4955 IU/ml，HBV为249 IU/ml，HAV为462 IU/ml，B19为460 IU/ml。我们目前方法的检测限为HCV 460 IU/ml，HIV-1 879 IU/ml，HBV 90 IU/ml，HAV 203 IU/ml，B19 314 IU/ml。
使用chemagic方法获得的95%检测限符合献血者筛查的监管要求。发现HCV、HBV、HAV和B19的检测灵敏度与手动纯化方法的灵敏度范围相似。然而，在本研究中发现chemagic方法对HIV-1的灵敏度较低。
