Journal
TRANSFUSION
Volume 40, Issue 6, Pages 718-724Publisher
AMER ASSOC BLOOD BANKS
DOI: 10.1046/j.1537-2995.2000.40060718.x
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BACKGROUND: High-throughput nucleic acid testing for transfusion-relevant viruses by PCR requires contamination-proof methods with high sensitivity and validity. A new PGR reagent kit (TaqMan, PE BioSystems) reduces the risk of carry-over contamination by eliminating post-PCR processing. STUDY DESIGN AND METHODS: Oligonucleotide design was done with software specialized for designing the assays' (TaqMan) primers and probes. A template-derived competitive internal control sequence designed through site-directed mutagenesis was used to reveal failures in amplification. Assay sensitivity was determined for single-donor and single-patient testing and by spiking sample mini-pools. Three seroconversion panels were tested. RESULTS: Sensitivity is high, reaching 300 HBV genomes per mt of single-patient material on direct testing. A detection limit of 1000 HBV genome equivalents per mi of donor plasma is achieved for 96 pooled samples. The window period for HBV infection was reduced by 17, 10, and 63 days from that for HBsAg screening in three seroconverting donors. CONCLUSION: The assay provides sufficient sensitivity to be superior to HBsAg screening in transfusion medicine and will be useful in clinical laboratories because of its ease of handling.
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