Single tube real-time RT-PCR Assay for the detection of enterovirus

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Publication Type Poster
School or College School of Medicine
Department Pathology
Program ARUP Institute for Clinical and Experimental Pathology
Creator Aldous, Wade K.; Hymas, Weston C.; Stevenson, Jeffery B.; Taggart, Edward W.;Hillyard, David R.
Title Single tube real-time RT-PCR Assay for the detection of enterovirus
Date 2004-09-11
Description We developed a rapid and sensitive method for the routine detection of enterovirus in CSF and plasma using a one tube, one step, real-time PCR assay. A primer set and an Eclipse probe were designed to target the highly conserved 5' untranslated region of the enterovirus genome. This assay was compared to a commercial PCR-ELISA assay. In an initial clinical validation, the newly developed real-time PCR assay provided 92% sensitivity, 100% specificity, 100% positive predictive value, and 97% negative predictive value compared to the commercial assay. During test validation, we detected 2 single nucleotide polymorphisms beneath the Eclipse probe sequence. To increase clinical sensitivity, we designed a modified degenerate Eclipse probe using neutral and specialized "super" bases from Epoch Biosciences (Bothell, WA). This modified probe assay demonstrated 100% agreement with the PCR-ELISA assay. The existence of unforeseen polymorphisms in genomes targeted by probes in realtime PCR assays can confound detection. Modified bases are useful tools for accommodating sequence variations in real-time assays.
Type Text
Publisher University of Utah
Subject MGB Eclipse; Viral RNA; Picornaviridae
Subject MESH Chemistry, Analytical; Clinical Laboratory Techniques; Immunoassay; Enterovirus Infections
Language eng
Bibliographic Citation Aldous WK, Hymas WC, Stevenson JB, Taggart EW, Hillyard DR. A Single Tube Real-Time RT-PCR Assay for the Detection of Enterovirus.
Format Medium application/pdf
Identifier ir-main,506
ARK ark:/87278/s6v41cmn
Setname ir_uspace
Date Created 2012-06-13
Date Modified 2012-06-13
ID 705548
Reference URL
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