Evaluation of an automated high-volume extraction method for viral nucleic acids in comparison to a manual procedure with preceding enrichment.
OBJECTIVE
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.


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Hourfar
M K
MK
Institute of Transfusion Medicine and Immunohaematology, German Red Cross, Johann Wolfgang Goethe University, Frankfurt, Germany.
Schmidt
M
M
Seifried
E
E
Roth
W K
WK
D001331:Automation; D001769:Blood / Q000821:virology; D001771:Blood Banks; D001782:Blood Donors; D001803:Blood Transfusion; D002498:Centrifugation; D004247:DNA / Q000032:analysis; D004279:DNA, Viral / Q000097:blood* / Q000302:isolation & purification*; D015497:HIV-1 / Q000235:genetics; D016174:Hepacivirus / Q000235:genetics; D030041:Hepatitis A virus / Q000235:genetics; D006515:Hepatitis B virus / Q000235:genetics; D006801:Humans; D008280:Magnetics; D021141:Nucleic Acid Amplification Techniques; D009696:Nucleic Acids / Q000302:isolation & purification; D016732:Parvovirus B19, Human / Q000235:genetics; D016133:Polymerase Chain Reaction; D012313:RNA / Q000032:analysis; D012367:RNA, Viral / Q000097:blood* / Q000302:isolation & purification*; D012680:Sensitivity and Specificity
D004279:DNA, Viral; D009696:Nucleic Acids; D012367:RNA, Viral; D012313:RNA; D004247:DNA