Development and characterization of new monoclonal antibodies against porcine Interleukin-17A and Interferon-gamma

Authors: MANIRARORA, JEAN - Food And Drug Administration(FDA); Walker, Kristen; PATIL, VEERUPAXAGOUDA - The Ohio State University; RENUKARADHYA, GOURAPURA - The Ohio State University; LABRESH, JOANNA - Kingfisher Biotech, Inc; SULLIVAN, YVONNE - Kingfisher Biotech, Inc; FRANCIS, ORE - University Of Bristol; Lunney, Joan

Submitted to: Frontiers in Immunology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/10/2022
Publication Date: 2/3/2022
Citation: Manirarora, J., Walker, K.E., Patil, V., Renukaradhya, G., Labresh, J., Sullivan, Y., Francis, O., Lunney, J.K. 2022. Development and characterization of new monoclonal antibodies against porcine Interleukin-17A and Interferon-gamma. Frontiers in Immunology. 13:786396. https://doi.org/10.3389/fimmu.2022.786396.
DOI: https://doi.org/10.3389/fimmu.2022.786396

Interpretive Summary: Pigs are a major food animal species in the US and worldwide. Losses caused by infectious diseases illustrate the need for effective diagnostics, therapeutics, and vaccine interventions. Unfortunately, there are limited commercial reagents to use to probe pig immunity, pathogen and vaccine responses. This manuscript describes the use of yeast expressed swine cytokines, interleukin-17A (IL-17A) and interferon gamma (IFNg), to generate panels of monoclonal antibodies (mAbs). These cytokines are important immune biomarkers, IL-17A as a regulator of host disease responses, and IFNg in anti-viral immunity. We used these mAbs to develop new assays for IL-17A and IFNg quantitation in swine fluids and for characterization of cytokine expressing cells. The availability of these new mAbs expands opportunities for swine immunology and biomedical research and will undoubtedly help dissect in more detail pathogen and vaccine responses in pigs.

Technical Abstract: Current research efforts require a broad range of immune reagents, but those available for pigs are limited. The goal of this study was to generate priority immune reagents for pigs and pipeline them for marketing. Our efforts were aimed at the expression of soluble swine cytokines and the production of panels of monoclonal antibodies (mAbs) to these proteins. Swine interleukin-17A (IL-17A) and Interferon-gamma (IFN') recombinant proteins were produced using yeast expression and used for monoclonal antibody (mAb) production resulting in panels of mAbs. All mAbs were checked for inhibition by other related mAbs to assign mAb epitopes. We also screened each new mAb for cross-species reactivity with orthologues of IL-17A or IFN'. Broad cross-reactions with dolphin, bovine, and ovine orthologues were found for several mAbs, as well as a unique cross-reaction identifying an anti-zebrafish IFN' mAb. For porcine IL-17A, a panel of 10 mAbs resulted in 8 different epitopes assigned, with the dolphin exhibiting the highest cross-species reactivities. For porcine IFN', a panel of 9 mAbs was characterized with 4 different epitopes; the highest cross-species reactions were found with dolphin, bovine, and caprine orthologues as well as a unique reaction of one mAb with zebrafish. The same panel of IL-17A mAbs was assessed to develop a quantitative sandwich ELISA detecting the yeast expressed protein as well as native IL-17A in stimulated peripheral blood mononuclear cell (PBMC) supernatants. The panel of anti-IL-17A mAbs was also screened for intracellular staining of stimulated PBMC and several identified as positive. These new mAbs expand opportunities for immunology research in swine.