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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
Author(s) -
Mitsunaga Shigeki,
Fujimura Kayoko,
Matsumoto Chieko,
Shiozawa Rieko,
Hirakawa Shinichi,
Nakajima Kazunori,
Tadokoro Kenji,
Juji Takeo
Publication year - 2002
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1046/j.1537-2995.2002.00024.x
Subject(s) - nucleic acid , virology , hepatitis b virus , real time polymerase chain reaction , dna , hepatitis c virus , rna , polymerase chain reaction , hepatitis b , biology , virus , gene , genetics
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. CONCLUSION: 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.