The detection of influenza A virus antigens in cultured cells by enzyme-linked immunosorbent assay.

An enzyme-linked immunosorbent assay (ELISA) was employed to investigate the expression of influenza A/Hong Kong/68 (H3N3) virus structural proteins on the surface of infected MDCK cells, and to detect viral antigens in culture media and cell extracts. Infected cells were fixed with 0.1 per cent glutaraldehyde before being examined for the presence of cell-surface antigens. Viral antigens were first observed on the surface of cells 4 hours after infection and reached a maximum 10-12 hours after infection, when measured by haemadsorption with chicken erythrocytes and by ELISA and immunofluorescence with hyperimmune antiserum to Hong Kong virus. A good correlation was found between the three assay systems. The presence of individual virion structural proteins on the cell surface was determined by ELISA using specific antibodies purified by differential affinity chromatography. Either or both or the internal matrix and nucleoprotein antigens were expressed from 2 to 6 hours after infection, with maximum expression after 2 hours, and the strain-specific and common antigenic determinants of haemagglutinin were observed on the cell surface from 4 hours after infection, and reached a maximum 8 to 10 hours after infection. Low levels of neuraminidase were detected between 4 and 8 hours after infection. Culture media and cell extracts were titrated by infectivity and haemagglutination assays, and by ELISA. Titres obtained from the culture media showed a close correlation between the three assay methods, with peak titres being attained 24 hours after infection. Viral antigens were first observed in cell extracts by ELISA 4 hours after infection, and infectious virions and haemagglutinin 2 hours later, but whereas maximum titres of infectious virus and haemagglutinin were found 10 hours after infection, the ELISA titre continued to rise until 24 hours after infection, which suggested that virus structural proteins were being accumulated in the cells after most of the progeny virions had been released. The results are discussed in terms of the potential use of ELISA in rapid virus diagnosis.