Nonspecific reactions in an enzyme-linked immunosorbent assay caused by binding of immunoglobulins in situ to egg-propagated infectious bronchitis virus.
High levels of nonspecific background absorbance and increased variability were found in a previously optimized enzyme-linked immunosorbent assay (ELISA) for infectious bronchitis virus (IBV) antibody after changing to commercially available non-pathogen-free eggs for viral antigen production. An increase in bound viral antigen in the assay caused a proportionate increase in the nonspecific binding of the conjugate, independent of other variables, in the absence of serum. Virus was propagated in non-pathogen-free eggs, and individual viral proteins were separated by polyacrylamide gel electrophoresis and transferred to nitrocellulose. Localization of chicken IgG-virus complexes were identified by immunoprecipitation with peroxidase-conjugated anti-chicken IgG. Specific staining at molecular weights corresponding to major proteins of IBV was demonstrated in these viral preparations. Virus grown in specific-pathogen-free eggs and treated in the same manner showed only slight amounts of staining. This evidence suggests that viral antigens grown in eggs from a non-pathogen-free flock bind with maternal chicken immunoglobulins present in the allantoic cavity of eggs. This IgG caused nonspecific reactions in our chicken ELISA system and gives cause for concern in any diagnostic system requiring the propagation of agents in fertile eggs.
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis*; D000956:Antigens, Viral / Q000378:metabolism*; D002642:Chick Embryo; D003332:Coronaviridae / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D007074:Immunoglobulin G / Q000378:metabolism*; D001351:Infectious bronchitis virus / Q000276:immunology* / Q000378:metabolism; D014776:Virus Cultivation
D000914:Antibodies, Viral; D000956:Antigens, Viral; D007074:Immunoglobulin G