A SIMPLE IMMUNOPEROXIDASE PLAQUE-ASSAY TO DETECT AND QUANTITATE MAREKS-DISEASE VIRUS PLAQUES

We report an immunoperoxidase-based staining technique that can be use d to rapidly and accurately detect and quantitate Marek's disease viru s (MDV) plaques. Monolayer cultures were fixed and incubated with a mo noclonal antibody specific for MDV. After washing, a second antibody o f horseradish peroxidase-conjugated goat anti-mouse IgG was applied, i ncubated for I hr, and washed with phosphate-buffered saline. After th e cultures were incubated with diaminobenzidine, CoCl2, and H2O2, the plaques appeared as black spots and were easily seen and counted. Sign ificantly more immunoperoxidase-stained serotype I MDV plaques could b e counted at 4 days postinoculation than were seen in unstained cultur es. With serotype 2 MDV-infected cells, the difference in plaque count s was less dramatic. Nevertheless, at 3 days postinoculation, signific antly more stained serotype 2 plaques were seen than unstained plaques . Immunoperoxidase staining of turkey herpesvirus plaques did not incr ease the sensitivity of viewing plaques. Similar numbers of stained an d unstained plaques were seen at 2 days postinoculation. We also demon strated that we could count serotype-specific MDV plaques in a mixed i nfection that contained all three serotypes.