Development of a competitive enzyme-linked immunosorbent assay for detection of bovine, ovine, porcine, and equine antibodies to vesicular stomatitis virus.

Two competitive (C) enzyme-linked immunosorbent assays (ELISAs) were developed for the detection of antibodies to vesicular stomatitis virus (VSV) in animal sera. The assays are based upon the availability of polyclonal antibodies (PAbs) from mouse ascitic fluids prepared against the New Jersey (NJ) and the Indiana (IN) VSV serotypes. The assays were performed by the immobilization of VSV-NJ and VSV-IN antigens on a solid phase (microtiter plate). Appropriately diluted test serum mixed with an equal volume of serotype-specific PAb was allowed to incubate in the presence of the relevant VSV antigen and finally with an enzyme-conjugated anti-mouse immunoglobulin. In the absence of anti-VSV antibodies in the test serum, the VSV antigen sites are reactive with the relevant PAb (NJ or IN) as indicated by color development after enzyme degradation of substrate. If the test serum contains the homologous VSV-NJ or VSV-IN antibodies, they compete with the relevant PAb for immobilized antigen sites and quantitatively block and inhibit the PAb reaction and subsequent color development. The performance of C-ELISAs in detecting anti-VSV antibodies in serum samples from four calves, two horses, four sheep, and seven pigs experimentally infected with VSV-NJ and VSV-IN was evaluated. The sensitivity and specificity of the C-ELISAs were compared with those of the standard microtiter serum neutralization (MTSN) tests. Homologous antibodies were demonstrable by C-ELISAs as early as 5 days postinfection (DPI) in one horse and one sheep infected with VSV-IN serotype. Seroconversion was demonstrable by C-ELISAs and MTSN tests in all animals by 9 DPI except in one sheep that received VSV-NJ and one horse inoculated with VSV-IN serotype which, on the basis of the MTSN test results, did not seroconvert until 14 and 11 DPI, respectively. The dynamics of homologous antibody response in all animals as revealed by the corresponding type-specific C-ELISAs paralleled the results of the MTSN tests. The type-specific antibodies to VSV serotypes increased exponentially during the first 2 to 4 weeks postinfection and remained relatively stable for about 6 months in some animals. The results suggest that the C-ELISAs offer many advantages over the MTSN tests and have potential applications as rapid and inexpensive tests in serodiagnosis of VSV infections in animals.

Simultaneous screening of bovine sera for antibodies to bluetongue and epizootic hemorrhagic disease of deer viruses by enzyme-linked immunosorbent assay.

An indirect enzyme-linked immunosorbent assay (I-ELISA) is described for simultaneous screening of bovine sera for detection of antibodies to bluetongue (BT) and epizootic hemorrhagic disease of deer (EHD) viruses (V). Optimal dilutions of BTV and EHDV antigens were combined and allowed to absorb on to the wells of microtiter plates. Appropriately diluted (1:100) bovine sera were allowed to incubate and the bound antibodies were detected by a murine monoclonal antibody (MAb) to bovine immunoglobulin (H-Chain) conjugated with horseradish peroxidase. The performance of the combined (C) I-ELISA in detecting antibodies to BTV and EHDV in sequential serum samples from calves experimentally inoculated with BTV, serotype 10, EHDV, serotype 1 (New Jersey) or EHDV serotype 2 (Alberta) was evaluated. Comparable antibody profiles were demonstrable by the CI-ELISA and separate I-ELISAs using either BTV or EHDV antigens. The results suggest that the CI-ELISA offers many advantages over the standard agar gel immunodiffusion (AGID) test and has potential application as a rapid, sensitive, inter-group-specific and inexpensive test for simultaneous screening of bovine sera for antibodies to BTV and/or EHDV.