Analytical specificity, sensitivity and variability testing:
Analytical specificity of the assay was determined using DNA extracted from cell culture amplified PRV strains Bartha, Becker, Shope, Bristol C61, and JS-2012, common swine viral pathogens encountered in modern swine herds, and bovine herpesvirus-1 (BHV-1), a related herpesvirus. For evaluating the intra-assay variability, 20 replicates of a single sample were tested in a single run. The inter-assay variability was calculated by running the same assay three times over 3 days. The analytical sensitivity of the triplex assay was determined using known concentration of PRV plasmid with net copy number of 7.34x1011. The limit of detection is interpreted as the lowest copy number that can be detected by the assay.