Development of dot-ELISA for the detection of venoms of major Indian venomous snakes

Abstract

India remained an epicenter for the snakebite-related mortality and morbidities due to widespread agricultural activities across the country and a considerable number of snakebites offended by Indian cobra (Naja naja), common krait (Bungarus caeruleus), Russell's viper (Daboia russelii), and saw-scaled viper (Echis carinatus). Presently, there is no selective test available for the detection of snake envenomation in India before the administration of snake antivenin. Therefore, the present study aimed to develop rapid, sensitive assay for the management of snakebite, which can detect venom, responsible snake species and serve as a tool for the reasonable administration of snake antivenin, which have scarcity across the world.The selective envenomation detection assay needs venom specific antibodies (VSAbs) for that monovalent antisera was prepared by hyperimmunization of rabbits with specific venom. However, obtained antibodies exhibit maximum activity towards homologous venom as well as quantifiable degree of cross-reactivity with heterologous venoms. Use of these antibodies for development of selective envenomation detection assay may create ambiguity in results, therefore needs to isolate VSAbs from monovalent antisera. The cross-reacting antibodies were specifically removed by immunoaffinity chromatography to obtain VSAbs. For the development of venom detection ELISA test (VDET), two different species of antibodies were used that offers enhanced sensitivity along with selective identification of the venoms of the responsible snakes.In conclusion, the developed VDET is rapid, specific, yet sensitive to detect venoms of offending snake species, and its venom concentration down to 1.0 ng/ml. However, the device observed with lowest venom concentration detection ability in the range <1.0 ng/ml from experimentally envenomated samples. The implementation of VDET will help in avoiding unnecessary usage and adverse reactions of snake antivenin. The test has all the merits to become a choice of method in envenomation diagnosis from medically important snakes of India.