Capture immunoassay for ruminant tumor necrosis factor-alpha: comparison with bioassay.
Monoclonal antibodies and IgG purified from rabbit polyclonal antiserum, raised against recombinant bovine tumor necrosis factor-alpha (TNF-alpha), have been employed in ELISA procedures to quantitate bovine TNF-alpha. These antibodies were potent in neutralizing the biological activity of recombinant as well as natural bovine TNF-alpha. The monoclonal antibodies were used as capture antibodies and were either passively adsorbed or covalently linked to ELISA plates. Polyclonal rabbit anti-TNF IgG was used as the detecting antibody in combination with a biotinylated anti-rabbit serum and a streptavidin-horseradish peroxidase conjugate. The detection limit for recombinant TNF-alpha medium was 10 pg ml-1 and in bovine or ovine serum was 35 pg ml-1. A good correlation was found between the ELISA and the WEHI-164 Clone 13 biologic assay when TNF-alpha was measured in medium containing serum or in serum. This capture ELISA was also capable of detecting ovine, but not porcine. TNF in supernatants from cultures of lipopolysaccharide-stimulated pulmonary alveolar macrophages.
Ellis
J A
JA
Veterinary Infectious Disease Organization (VIDO), Saskatoon, Sask, Canada.
Godson
D
D
Campos
M
M
Sileghem
M
M
Babiuk
L A
LA
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001681:Biological Assay / Q000379:methods* / Q000662:veterinary; D002417:Cattle; D002478:Cells, Cultured; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods* / Q000662:veterinary; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000276:immunology; D016676:Macrophages, Alveolar / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D011817:Rabbits; D011994:Recombinant Proteins / Q000032:analysis; D012418:Ruminants / Q000276:immunology*; D012680:Sensitivity and Specificity; D012756:Sheep; D013552:Swine; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis*
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha