Detection of anti-TNFa activity in canine hyperimmune serum using a TNFa inhibition assay

Kotiw, Michael and Morgan, Michael and Taylor, Stephen M. and Shiels, Ian A. (2010) Detection of anti-TNFa activity in canine hyperimmune serum using a TNFa inhibition assay. Veterinary Clinical Pathology, 39 (1). pp. 46-52. ISSN 0275-6382


Background: Increased serum tumor necrosis factor-α (TNFα) activity has been associated with onset of serious inflammatory diseases in dogs. Development of treatment with TNFα-antagonists has been limited by the unavailability of suitable reagents and potency assays for TNFα. Objectives: The objectives of this study were to optimize a cell-based assay to measure anti-TNFα activity in serum and plasma from hyperimmune (vaccinated with an Escherichia coli J5 bacterin) and unvaccinated canine donors; to use the assay to determine whether hyperimmune serum inhibits TNFα activity in vivo; and to determine whether soluble TNF receptor-1 (sTNFR1, a naturally occurring TNFα antagonist) contributes to anti-TNFα activity. Methods: Commercial plasma and serum from hyperimmune-frozen plasma (HFP) donors and unvaccinated fresh-frozen plasma (FFP) donors were used in the study. An L929-cell TNFα-inhibition assay (LTIA) was optimized to measure anti-TNFα activity. Using a rat subcutaneous pouch model of inflammation, the effects of HFP, FFP, a synthetic TNFα antagonist (Etanercept), and carprofen on TNFα activity were compared in vivo. Immunofluorescence was used to measure soluble sTNFR1 concentration. Results: Using the optimized LTIA, HFP serum but not FFP serum decreased canine TNFα activity (P<.01). HFP plasma and Etanercept (but not FFP plasma or carprofen) significantly decreased TNFα activity in pouch exudates (P<.05). A significantly higher concentration of sTNFR1 was found in HFP than FFP serum. Conclusions: Using the LTIA, anti-TNFα activity is readily measured in canine serum and inflammatory exudates. sTNFR1 appears to contribute to anti-TNFα activity in HFP serum. These results suggest HFP should be investigated further as a potential immunotherapeutic agent for controlling canine diseases in which TNFα is implicated.

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Item Type: Article (Commonwealth Reporting Category C)
Refereed: Yes
Item Status: Live Archive
Additional Information: Permanent restricted access to Published version due to publisher's copyright restrictions.
Faculty / Department / School: Historic - Faculty of Sciences - Department of Biological and Physical Sciences
Date Deposited: 08 Dec 2010 11:22
Last Modified: 02 Jul 2013 23:57
Uncontrolled Keywords: dogs; escherichia coli J5; hyperimmune serum; L929 cell assay; TNF inhibition
Fields of Research : 11 Medical and Health Sciences > 1107 Immunology > 110702 Applied Immunology (incl. Antibody Engineering, Xenotransplantation and T-cell Therapies)
06 Biological Sciences > 0605 Microbiology > 060501 Bacteriology
07 Agricultural and Veterinary Sciences > 0707 Veterinary Sciences > 070705 Veterinary Immunology
Socio-Economic Objective: E Expanding Knowledge > 97 Expanding Knowledge > 970111 Expanding Knowledge in the Medical and Health Sciences
Identification Number or DOI: 10.1111/j.1939-165X.2009.00166.x

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