Evaluation of a solid-phase immunoassay with fluorescein isothiocyanate-conjugated heterogeneous or monoclonal antibodies for identification of virus isolates, with influenza virus as a model

Abstract

A solid-phase immunofluorescence assay was evaluated for the identification of viruses isolated in tissue culture, with influenza virus as a model. Purified immunoglobulin G (IgG) from hyperimmune rabbit sera specific for contemporary strains of influenza A or B was covalently attached to microscopic plastic beads to capture virus. Fluorescein isothiocyanate (FITC)-conjugated antibodies of different specificities were then reacted with bound antigen, and the resulting complexes were quantified in a suitable filter fluorometer. The assay, with appropriately absorbed FITC-conjugated second antibody, reliably identified virus present in harvests from cell cultures infected with clinical specimens. For influenza A (H1N1) virus, sensitivity of detecting antigen was about 8- to 32-fold less when an FITC-conjugated monoclonal Igg antibody pool specific for epitopes in three different antigenic sites on influenza hemagglutinin was used as the second antibody as compared to when IgG from hyperimmune sera specific for virus or its components was used as the second antibody. The immunofluorometric assay provides a method for quantitative detection of viral antigen in tissue culture fluids and objective identification of virus type and subtype with FITC-conjugated reagents.