Abstract
Eight laboratories worked collectively to evaluate 4 real-time RT-PCR (rRT-PCR) protocols targeting viral hemorrhagic septicemia virus (VHSV) being considered for deployment to a USA laboratory testing network. The protocols utilized previously published primers and probe sets developed for detection and surveillance of VHSV. All participating laboratories received and followed a standard operating protocol for extraction and for each of the rRT-PCR assays. Performance measures specifically evaluated included limit of detection (defined as the smallest amount of analyte in which 95% of the samples are classified as positive), analytical specificity, assay efficiency across genotype representatives, within- and between-plate variation within a laboratory, and variation between laboratories using the same platform, between platforms, and between software versions. This evaluation clearly demonstrated that the TaqMan®-based assay developed by Jonstrup et al. (2013; J Fish Dis 36:9-23) produced the most consistent analytical performance characteristics for detecting all genotypes of VHSV across the 8 participating laboratories.
Original language | English (US) |
---|---|
Pages (from-to) | 1-13 |
Number of pages | 13 |
Journal | Diseases of aquatic organisms |
Volume | 111 |
Issue number | 1 |
DOIs | |
State | Published - Aug 21 2014 |
Keywords
- Analytical sensitivity
- Analytical specificity
- Real-time RT-PCR
- Surveillance
- VHSV
- Validation