An immunoperoxidase plaque assay for reticuloendotheliosis virus and its application to a sensitive serum neutralization assay.
A rapid assay for the enumeration of reticuloendotheliosis virus (REV) is described. Chicken embryo fibroblast monolayer cultures were infected with REV and incubated 6 days under an agar overlay. After removal of the overlay, cells were fixed with acetone/ethanol. Foci of infection (hereafter referred to as plaques) were detected using either an anti-REV envelope monoclonal antibody or convalescent chicken serum as the primary antibody; the secondary antibody was either horseradish peroxidase-conjugated goat anti-mouse IgG (for use with monoclonals) or goat anti-chicken IgG (for use with chicken serum). Staining with a substrate solution containing diaminobenzidine, CoCl2, and H2O2 revealed individual dark plaques on a light gray background. This method worked equally well for the SNV, CSV, and REV-T strains of REV; further, it detected all six field isolates tested. Results indicate that this immunoperoxidase technique is a rapid and reliable method for detection and titration of REV as well as for the assay of neutralizing antibody in chicken serum.
Calvert
J G
JG
U.S. Department of Agriculture, Agricultural Research Service, Avian Disease and Oncology Laboratory, East Lansing, Michigan 48823.
Nazerian
K
K
D000818:Animals; D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D002478:Cells, Cultured; D002642:Chick Embryo; D002645:Chickens; D005347:Fibroblasts; D006041:Goats / Q000276:immunology; D007124:Immunoenzyme Techniques; D051379:Mice / Q000276:immunology; D009500:Neutralization Tests / Q000662:veterinary*; D011201:Poultry Diseases; D001356:Reticuloendotheliosis virus / Q000302:isolation & purification*; D012192:Retroviridae Infections / Q000175:diagnosis / Q000662:veterinary*; D014412:Tumor Virus Infections / Q000175:diagnosis / Q000662:veterinary*; D014759:Viral Envelope Proteins / Q000032:analysis; D010948:Viral Plaque Assay
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D014759:Viral Envelope Proteins