Development of anti-bovine TNF-alpha mAb and ELISA for quantitating TNF-alpha in milk after intramammary injection of endotoxin.
Murine mAb reactive with recombinant bovine tumor necrosis factor-alpha (r-boTNF-alpha) were produced. An ELISA using murine mAb and rabbit polyclonal antibodies, each reactive with r-boTNF-alpha to sandwich bovine TNF-alpha was developed. Secretion of TNF-alpha in quarter milk increased 1 h after injection of 0.1 mg (four cows) or 0.5 mg (four cows) Escherichia coli lipopolysaccharide (LPS) into a mammary quarter, peaked 1 to 5 h later, and returned to control levels in 24 h. There were no differences in body temperature, SCC, TNF-alpha, and blood leukocyte responses between 0.1 and 0.5 mg of LPS. To determine effects of repeated injections of LPS into the same udder, a second injection of 0.1 mg of LPS into the same quarter (two cows) 24 h after the first injection produced a strongly attenuated TNF-alpha response. However, a normal TNF-alpha response was observed when LPS was injected into a contralateral quarter (two cows) 24 h after the first LPS injection. Leukocyte counts in blood decreased and body temperature increased substantially after each injection of LPS. Quarter milk SCC increased 200-fold 8 to 12 h after the LPS injections. It would appear that these changes were not regulated by TNF-alpha secretion because the changes were also similar after the second injection of LPS into the same mammary quarter.
Paape
M J
MJ
Immunology and Disease Resistance Laboratory, ARS, USDA, Beltsville, Maryland 20705, USA. mpaape@anri.barc.usda.gov
Rautiainen
P M
PM
Lilius
E M
EM
Malstrom
C E
CE
Elsasser
T H
TH
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000276:immunology; D001831:Body Temperature / Q000187:drug effects; D002417:Cattle / Q000276:immunology / Q000378:metabolism*; D004306:Dose-Response Relationship, Immunologic; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D004926:Escherichia coli; D005260:Female; D007700:Kinetics; D007958:Leukocyte Count / Q000662:veterinary; D008070:Lipopolysaccharides / Q000276:immunology / Q000494:pharmacology*; D008321:Mammary Glands, Animal / Q000166:cytology / Q000187:drug effects / Q000276:immunology; D051379:Mice; D008892:Milk / Q000166:cytology / Q000276:immunology*; D011817:Rabbits; D014409:Tumor Necrosis Factor-alpha / Q000032:analysis* / Q000276:immunology
D000911:Antibodies, Monoclonal; D008070:Lipopolysaccharides; D014409:Tumor Necrosis Factor-alpha