Baculovirus expression of the fusion protein gene of bovine respiratory syncytial virus and utility of the recombinant protein in a diagnostic enzyme immunoassay

Abstract

The fusion (F) protein of bovine respiratory syncytial virus (BRSV) was expressed by using a baculovirus vector. Antigenicity was tested by immunofluorescence analysis with F-specific monoclonal and polyclonal antibodies. Antibodies to recombinant F protein raised in a rabbit neutralized BRSV and human respiratory syncytial virus infectivity when tested in a plaque reduction assay. The recombinant F protein was evaluated as a source of antigen in an enzyme-linked immunosorbent assay (ELISA), and this ELISA was compared with the virus neutralization (VN) test for detecting BRSV antibodies in 10 consecutive serum samples from four calves vaccinated with a live modified BRSV vaccine and from two nonvaccinated control calves. The ELISA with the baculovirus-expressed F protein as an antigen compared favorably with the VN test and is a rapid, sensitive, and specific method for detecting serum antibodies to BRSV.

FIG. 1

Expression of F protein by recombinant baculovirus and confirmation of the authenticity of recombinant F protein. Lanes: A, molecular weight standards; B, recombinant baculovirus-infected cell lysate immunoprecipitated with BRSV-specific antiserum; C, BRSV-infected BTu cell lysate immunoprecipitated with BRSV antiserum; D, recombinant baculovirus-infected cell lysate immunoprecipitated with baculovirus-expressed F-specific antiserum; E, recombinant baculovirus-infected cell lysate; F, BRSV-infected BTu cell lysate; G, wild-type AcNPV-infected cell lysate; H, molecular weight standards. Numbers are molecular weights, in thousands.

FIG. 2

ELISA and VN test results obtained with serum samples from four vaccinated calves. Calves were inoculated intramuscularly on day 0 with an attenuated BRSV vaccine, and a booster vaccination was given after 4 weeks. The ELISA and the VN test were performed as described in the text.