ONE ELISA FOR ALL ARTIODACTYLIDS: ASSESSMENT OF A PROTEIN G CONJUGATE FOR DETECTION OF ANTIBODY IN NONDOMESTIC HOOFSTOCK

摘要

The Johne's disease serum antibody enzyme-linked immunosorbent assay (ELIS A) has been a useful herd infection screening tool for domestic agriculture species. This protein-G conjugate, test provides valuable diagnostic information and is both fast and inexpensive. Since the ELISA assay has been validated only for cattle in the United States, screening for M. paratuberculosis infection in U.S. captive and free-ranging non-domestic species has not benefitted from ELISA technology. Johne's disease surveillance thus has been limited in most cases to fecal culture. The ability of protein G to bind immunoglobulin in well-characterized laboratory species (rat, mouse, human, etc.) has been shown, but its ability to bind immunoglobulin from most non-domestic hoofstock species has not been well described. To determine if this antibody detection system can be used effectively for multiple non-domestic species, the binding characteristics of the protein G conjugate was assessed for 12 non-domestic hoofstock species and two domestic control species (bovine [positive control], chicken [negative control]; elk, muntjac, white tail deer; bison, sheep, bontebok, impala, llama, kudu/nyala, addax, antelope, oryx). Partially purified immunoglobulin was obtained through high performance liquid chromatography (HPLC) of sera from six to eight healthy adult individuals belonging to each of the 14 species. Two methods (size-exclusion chromatography and SDS-PAGE) were used to confirm the presence of antibody and to quantify recovery. The binding characteristics of the protein G-horseradish peroxidase detection system utilized in the ELISA were compared for each sample. This was accomplished by measuring the optical density of a uniform amount of immunglobulin from each of the 106 individual samples when tested against seven concentrations (2.0-5.0 g/ml) of solid phase protein G (the antigen).