Optimization and validation of ELISAs for interferon-gamma determination in bison.

Abstract

Bovine tuberculosis, caused by Mycobacterium bovis, is endemic in the Wood Buffalo National Park, Canada, home to free-ranging and threatened wood bison. This disease poses a threat to the conservation of this culturally and ecologically important animal species, as well as potentially impacting the health of humans and other animal species via zoonosis and spillover, respectively. The ability to detect infection early will minimize and prevent the potential risk of M. bovis transmission. Interferon-gamma (IFNγ) assays are a reliable detection method for M. bovis in cattle and other wildlife species and may have diagnostic value in bison as well. We aimed to optimize and partially validate 2 commercial IFNγ ELISAs to detect endogenous bison IFNγ in mitogen-stimulated whole blood. Parameters evaluated included antibody identification, sample matrix effect, dilution linearity, assay reproducibility, and limit of quantification. The optimized assays demonstrated linear responses to recombinant bovine and endogenous bison IFNγ (range: 1-125 pg/mL; R² = 0.99), with good recovery and fair reproducibility, and a low limit of quantification of 1 pg/mL. Mabtech bovine Flex and Pro kits have the same antibodies but in 2 different assay formats; an in-house assay platform (Flex kit) and precoated plates (Pro kit) are considered suitable for measuring bison IFNγ, offering flexibility depending on available resources.