Critical aspects on traditional antivenom production processes and their optimization by factorial analysis.

Most antivenoms are produced by techniques developed over 50 years ago, with minor modifications. Herein we revise the core of traditional antivenom production processes aiming to optimize key determinants for both consistent antivenom production and the best balance between F(ab')2 quality and recovery. Factorial design analysis revealed that pepsin digestion of 1:3 saline diluted equine plasma for 60 min under pH: 3.20, 37 °C temperature and a 1:15 pepsin to protein ratio conditions, allowed to achieve maximal IgG to F(ab')2 conversion with minimal protein aggregate formation. Further downstream processing by salting out with ammonium sulfate was also studied by factorial analysis. The influence of ammonium sulfate (AS) concentration, temperature (T) and the albumin to total plasma protein ratio plasma (Alb:P) were assayed, revealing that both AS, T and their interaction have a significant impact in F(ab')2 quality and recovery. Taking into account the existing compromise between F(ab')2 monomer recovery and quality two alternative conditions were selected: 14 g/dl AS at 56 °C and, alternatively 16 g/dl AS at 30 °C. Reasonable yields (42%) and product quality (2.5% of aggregates) without significant changes in production cost of traditional methodologies was achieved under the optimized conditions found.

Effectiveness of two common antivenoms for North, Central, and South American Micrurus envenomations.

Micrurus snakes (coral snakes) may produce severe envenomation that can lead to death by peripheral respiratory paralysis. Only few laboratories produce specific antivenoms, and despite the cross-reactivity found in some Micrurus species venoms, the treatment is not always effective. To test two therapeutic antivenoms against the venom of four species of Micrurus from Southern America, North of South America, Central America, and North America, the determination of the lethal potency of the venoms, the study of some biochemical and immunochemical characteristics, and the determination of the neutralizing activity of both antivenoms were studied. North American and South American antivenoms neutralized well venoms from Micrurus species of the corresponding hemisphere but displayed lower effectiveness against venoms of species from different hemispheres. It was concluded that the neutralization of Micrurus venoms by regional antivenoms could be useful to treat the envenomation by some Micrurus snakes but is necessary to evaluate carefully the antivenoms to be used with the venoms from the snakes of the region. Also, considering the difficulties for coral snake antivenom production, the development of a polyvalent antivenom is useful to treat the envenomation by coral snakes from different regions is necessary.