Production of an alternative antivenom based on egg yolk antibodies against Crotalus durissus terrificus

CANGELOSI, A.; LEIVA C.; MARICONDA V.; BRERO ML; CHACANA P.; GEOGHEGAN, PATRICIA

Buenos Aires

Congreso; 20th World Congress of the International Society on Toxinology; 2019

Snakebite envenoming represents an example of a pathology which effective treatment is the administration of antivenoms. An alternative to mammal polyclonal sera is the use of egg yolk antibodies due to its advantages regarding animal welfare and lower costs of production. The aim if this study is to produce an IgY antivenom against Crotalus durissus terrificus at a pilot scale using more simplified methods of purification. A group of laying hens (n=2) were immunized via i.m. with 80 μg of C. d. terrificus whole venom (pool of venoms of snakes from Argentina) nine times at days 0, 14, 28, 71, 237, 289, 304, 473 and 487. For the first immunization, venom was emulsified with Freund`s complete (1st injection) and incomplete adjuvants (boosters). Eggs were collected during 10 days after the 8th and 9th immunization. In order to choose the optimal method of purification, different methods of purification were evaluated: ammonium sulphate precipitation (24 and 26% w/v), PEG-6000 (12% w/v) and caprylic acid (7% v/v). 0.01% (w/v) thimerosal was added for preservation. Median effective dose (ED50) was assessed in mice by mixing 3 LD50 (17 μg) with increasing volumes of IgY antivenom according to WHO guidelines (World Health Organization, 2017). The optimal ED50 was obtained by sulphate ammonium precipitation rather than using PEG-8000 and caprylic acid. After 9 immunizations, 1 ml IgY antivenom purified by PEG-8000 neutralized 158 μg of venom. In addition, 1 ml IgY antivenom purified by caprylic acid neutralized < 40 μg of venom. However, 1 ml IgY antivenom purified by sulphate ammonium neutralized 395 μg of venom. In conclusion, immunization of hens with sub-lethal doses of C. d. terrificus venom produced antivenoms with DE50 similar to the obtained in horses. Thus, IgY-technology may allow the production of effective and affordable antivenoms.