PCR and qPCR
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Double-quenched probes increase sensitivity of qPCR assay detecting viral load

Muldrew K, Lovett J. (2013). An In-House Assay for BK Polyomavirus Quantification Using the Abbott m2000 RealTime System. J Med Microbio, 17141720. PMID: 23924663.

Viral load of BK polyomavirus (BKV) is a useful indicator of BKV disease and renal transplant patient response to therapy. Here the authors describe their development of a fast, sensitive, and cost-effective, in-house qPCR assay targeting the BK polyomavirus major capsid VP1 gene for monitoring viral load.

As part of assay development, the researchers evaluated the use of ZEN™ Double-Quenched Probes (included as part of IDT PrimeTime® qPCR Assays), which demonstrated a marked decrease in background fluorescence compared to an identical TaqMan® probe containing only a single quencher, and they suggest that such double-quenched probes may be a better approach for other qPCR probe-based assays. Serial low level dilutions demonstrated increased sensitivity of the probe containing the ZEN Quencher, resulting in an assay with a wider linear range of detection. The assay protocol was developed specifically for the Abbott m2000 RealTime System to increase throughput, and successfully decreased handling and overall assay time, as well as cost per reaction compared to similar BKV assays performed in other clinical settings.


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