Evaluation of a Monoclonal Antibody Blocking ELISA for the Detection of Group-specific Antibodies to Bluetongue Virus in Experimental and Field Sera

Abstract

SUMMARYIn order to overcome serological cross-reactions among orbivirus serogroups, which can hinder the accurate diagnosis of bluetongue virus (BTV) infection of livestock, a blocking ELISA (B-ELISA) incorporating a monoclonal antibody (20E9B7G2) with specificity for the BTV serogroup was developed. Experimental antisera raised to South African BTV serotypes 1 to 19 were tested in the B-ELISA and all blocked the binding of 20E9B7G2 to BTV antigen. The sensitivity and specificity of the assay was evaluated with a range of experimental and field sera and compared to a sensitive indirect ELISA (I-ELISA) for the detection of BTV-specific antibodies. The specificity of the B-ELISA was absolute for antibodies to BTV, showing no cross-reaction with experimental antisera to serotypes of the closely related orbivirus causing epizootic haemorrhagic disease of deer. The sensitivity of the B-ELISA exceeded that of the I-ELISA. In particular, the B-ELISA detected a BTV-specific antibody response much earlier after infection that the I-ELISA, while still exhibiting full sensitivity to BTV antibody titres several months after infection.

Detection of PMV‐1 specific antibodies with a monoclonal antibody blocking enzyme‐linked immunosorbent assay

“…By selecting the right Mab, this test can be adapted either for differentiation between serotypes (Anderson, 1984;Lunt et al, 1988) or for the detection of antibodies to well-conserved, species-specific epitopes (Belinda et al, 1995;Czifra et al, 1995).…”

Major core protein VP7 of Australian bluetongue virus serotype 15: sequence and antigenicity divergence from other BTV serotypes

“…Sera from four mice immunized with bacterial recombinant BTV-15 VP7 were tested for their reactivity using purified BTV-15AU and BTV-1AU virus as antigens. Preparation and quantification of viral antigens and indirect ELISAs were carried out as described by Lunt et al (1988). Sera to be tested were serially diluted fourfold with a starting dilution of 1:200.…”

“…More recently it was found that most, if not all, anti-VP7 monoclonal antibodies (MAbs) raised against Australian BTV-1 failed to react with Australian BTV-15 antigens (White, 1992). Competition ELISA (C-ELISA) using either viral VP7 or recombinant VP7 protein, and VP7-specific MAbs, have been developed and shown to be highly effective in the specific detection of anti-BTV antibodies in sera (Lunt et al, 1988;Martyn et al, 1990;Anderson, 1984;Afshar et al, 1987Afshar et al, , 1992. However a number of these tests suffer from a relatively low efficiency of detection of anti-BTV-15 antibodies.…”

Conformation of the VP2 protein of bluetongue virus (BTV) determines the involvement in virus neutralization of highly conserved epitopes within the BTV serogroup

“…MAbs were screened by ELISA as previously described (Lunt et al, 1988b). The hybridoma secreting 9J3/G6 was isolated following fusion of SP2/0-Agl4 cells with spleen cells (Breschkin et al, 1981) from mice inoculated with a 10 K suspension of BTV-1-infected suckling mouse brain in Hanks' balanced salt solution.…”