Rapid, label-free surface plasmon resonance discrimination between Bothrops and Crotalus venoms using clinical antivenom as the capture reagent

Snake envenoming is recognized as a global health problem, affecting thousands of people every year. One of the main challenges in addressing this issue is the correct identification and treatment of these envenomations, mainly in locations where people don’t have easy access to hospitals. In Brazil, the genus Bothrops is responsible for the majority of envenomations, followed by Crotalus. This study reports a simple methodology for detecting raw venom from snakes of the genus Bothrops, through interaction with their corresponding antibodies, using a high-sensitivity optical biosensor. The protocol consists of adding antibodies (present in the commercial antivenom) to the sensor surface, followed by the addition of Crotalus venom (nonspecific), and then Bothrops venom (specific), resulting in changes in the refractive index, to evaluate cross reactions between them. Different concentrations of raw venom from snakes of the genus Bothrops and Crotalus were tested, starting with a concentration of 6.784μgmL ⁻¹ and progressing until reaching the minimum detectable concentration. The binding capacity of venom to antivenom was investigated at two concentrations of antivenom: 5 μgm ⁻¹ and 50 μgmL ⁻¹ . Both antivenom and snake venoms were solubilized in phosphate buffered saline (PBS). The results show an accurate detection of the antigen of interest (Bothrops venom), tested at different concentrations. The biosensor was able to detect venom up to a concentration of 0.848 μgmL ⁻¹ . In addition, no interference from nonspecific binding between the Bothrops antivenom and Crotalus venom was detected. The detection of specific venom (Bothrops) occurred in a satisfactory time (up to 14 minutes). The results provide evidence that the biosensor and the methodology employed can be considered a diagnostic model under development, which can help health workers to better identify and treat envenomated people.