Production and assessment of ovine antisera for the manufacture of a veterinary adder antivenom

Medically important venomous snakes in Western Europe are Vipera ammodytes, Vipera aspis, Vipera berus and Vipera latastei. Envenomation of dogs and other animals by these snakes receives limited attention despite the relative frequency and potential mortality and morbidity. This reflects, in part, the lack of a dedicated veterinary antivenom. Successful antivenoms are derived from antisera containing high levels of specific polyclonal antibodies that bind to, and neutralise, all the toxins present. This requires a careful choice of immunogen, animals and immunisation schedule. We detected proteomic variation in the venoms of V ammodytes, V aspis, V berus and V latastei by SDS-PAGE (sodium dodecyl sulphate-polyacrylamide gel electrophoresis) gel electrophoresis. Consequently, we used a mixture containing equal amounts of venom from these species to immunise a flock of sheep. We demonstrate that immunisation resulted in antisera containing high levels of specific antibodies directed against the majority of toxic components found in all four snake venoms using immunoblotting, ELISA and small-scale affinity chromatography assays. The latter shows that all 25 sheep responded quickly and maintained high levels of specific antibodies throughout the two-year period of study. This ensures a consistent starting material for the manufacture of a reproducible veterinary antivenom, ViperaVet. Our next objectives are to purify the antibodies from our antisera and demonstrate their preclinical neutralising efficacy in murine animal studies prior to undertaking a clinical trial in envenomed patients.