Use of venom blocked with chelating agent for anti-bothropic serum production

LOPEZ GISELA; VAN DE VELDE, A; DAVID HERNANDEZ; LAURA LEIVA; FUSCO, L

Congreso; Venoms and Toxins 2020 | Virtual; 2020

OXFORD

Snakes belonging to genus Bothrops are responsible for more than 85% of the bites occurring inSouth America. Bothrops alternatus is a pitviper widespread in this area and it is one of the mostimportant species associated to snakebites not only in Argentina but also Brazil. Antivenoms arethe only specific treatment for envenoming by snakebites. They are made by animals (e.g. horses orsheep) immunizations with sublethal doses of venom. Snake Venom Metalloproteinases (SVMPs)play an important role in envenomation, causing relevant local effects such as hemorrhage, edemaand myotoxicity as well as systemic bleeding. They represent around ̴43.1% of the proteincomposition from B. alternatus venom and causing lesions during the immunization of animals. Inthis work, an alternative immunization protocol was developed in mice where SVMPs activity waspreviously blocked by Na2EDTA as chelating agent. For this proposal, the B. alternatus venom (BaV,10 mg/mL) was treated with 50 mM Na2EDTA (1 h, 37°C) and excess chelator wasremoved by Sephadex G-25 chromatography. Proteolytic activity was assayed to control the blockprocessing. Groups of 5 BALB/c mice were immunized s.v. on 0-15-30 days with BaV (15-30-45μg)and BaV/Na2EDTA (45-90-135 μg). Blood samples were collected on days 14-29-41 for antibodyanalysis. Sera from BaV/Na2EDTA protocol have a titer (5.1x104) higher than those treated with BaV(1.3x104). The neutralizing ability of both antivenoms was tested against proteolytic, coagulant andPLA2 activity, resulting that it was significantly higher (p