Development of an innovative methodology of antivenomics approach, combining the use of magnetic beads and mass spectrometry.

Snakebite is classified as a Category A neglected tropical disease by the WHO, as it causes the
death of about 150,000 people every year, mostly in rural and poor areas of the world. Snake
envenoming is classically treated by injecting antivenom, which is antitoxin antibodies (Igs)
collected from immunized animals. However, these treatments may induce immunological
reactions including severe adverse reaction to the patient. Moreover, venom compositions
strongly differ from species, gender and habitat, explaining why providing broadly effective
antivenoms is a real challenge. In this context, quantitatively evaluating the toxin-binding
capability of any antivenom is crucial to improve the production of effective snakebite
therapeutics. In this study, we propose an alternative methodology for the so-called
“antivenomics” methodology. Indeed, affinity columns coupled to mass spectrometry have
been demonstrated performant, but their preparation and their lifetime represent constraints
to the throughput of antivenom evaluation. In this work, we exploit the potential of magnetic
beads, LC-MS and shotgun proteomics mass spectrometry to speed up antivenom efficacy
characterization. The antivenom Igs are bound to magnetic beads, before being incubated in
the presence of various venoms. Comparative MS analysis of the toxins remaining in
suspension (not recognized by Ig) and those remaining on the beads (recognized by Ig) allows
the binding selectivity of the antivenom to be determined. The strategy is demonstrated here
with venom from the medically important African viper, Echis ocellatus.