Factors affecting virus plaque confirmation procedures

Direct plaque counts obtained by using the monolayer cell culture assay technique reliably confirmed the number of viruses isolated. Analysis revealed some significant differences in the false-positive rate, depending on the test method used or virus samples evaluated. Plaques from laboratory stock viruses showed a higher confirmation rate than sewage plaque isolates. Test results with laboratory stock viruses suggested that confirmation rates may be affected by virus types present in the sample. Plaques picked by the stab and scrape method and immediately passed into cell culture tubes produced the most reliable counts as compared to those picked by the stab only method or those stored at -70 degrees C in Earle's balanced salt solution with or without fetal calf serum. Plaque confirmation using this method was 90% or better. Although the term 'false positive plaque' has been applied to a particular plaque that was not confirmed, five of ten plaques picked by the stab and scrape method in one series of experiments were confirmed when repicked by the same method from original plaque bottles, indicating that a substantial number of unconfirmed plaques may be caused by plaque transfer techniques.