High-throughput HBV DNA and HCV RNA detection system using a nucleic acid purification robot and real-time detection PCR: its application to analysis of posttransfusion hepatitis.
BACKGROUND
A high-throughput detection system was developed for HBV DNA and HCV RNA.


METHODS
A combination of real-time detection PCR using an automated system (PRISM 7700, PE Biosystems, Foster City, CA) and automatic viral nucleic acid extraction (BioRobot 9604, Qiagen, Hilden, Germany) was used as the high-throughput detection system. An internal control for HBV DNA detection was also developed.


RESULTS
Testing of 96 samples for HBV and HCV was completed within 5 hours. The sensitivity of this system almost equals that of the manual method using nested PCR. The addition of an internal control for HBV detection did not affect the sensitivity of the method and confirmed the accuracy of results. It was possible to quantify HBV in HBV+ samples that contain more than 500 genome equivalents per mL. We started using this system from June 1999 for testing stored donor and patient samples to analyze cases of posttransfusion hepatitis and identified three HBV+ donations that were implicated in posttransfusion hepatitis B.


CONCLUSIONS
The high-throughput detection system is a useful tool for HBV DNA and HCV RNA detection because it enables rapid and reliable testing of a large number of samples.
Mitsunaga
Shigeki
S
Transfusion Information Department, The Japanese Red Cross Central Blood Center, Tokyo, Japan. mitsu@cbc.jrc.or.jp
Fujimura
Kayoko
K
Matsumoto
Chieko
C
Shiozawa
Rieko
R
Hirakawa
Shinichi
S
Nakajima
Kazunori
K
Tadokoro
Kenji
K
Juji
Takeo
T
D001331:Automation; D003199:Computer Systems; D004279:DNA, Viral / Q000097:blood*; D016174:Hepacivirus / Q000235:genetics / Q000302:isolation & purification*; D006509:Hepatitis B / Q000097:blood* / Q000175:diagnosis / Q000635:transmission / Q000821:virology; D006515:Hepatitis B virus / Q000235:genetics / Q000302:isolation & purification*; D006526:Hepatitis C / Q000097:blood* / Q000175:diagnosis / Q000635:transmission / Q000821:virology; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D012367:RNA, Viral / Q000097:blood*; D015203:Reproducibility of Results; D012371:Robotics; D012680:Sensitivity and Specificity; D065227:Transfusion Reaction; D014766:Viremia / Q000097:blood* / Q000175:diagnosis / Q000635:transmission / Q000821:virology
D004279:DNA, Viral; D012367:RNA, Viral