Comparison of Antivenom potential of chicken Egg yolk Antibodies Generated against Bentonite and Adjuvant coated Echis carinatus venom

摘要

The Chicken egg yolk antibodies generated against Bentonite and Adjuvant (FCA) coated Echis carinatus venom was evaluated for their anti-venom potential. Antibodies are extracted from immunized chicken egg yolk by the method described by Polson et al.(1980) and further purified by DEAE cellulose ion exchange column chromatography, which gave pure (180 - 200KDa) specific antibodies against venom. Inhibition of lethal, edema, procoagulant, and phospholipase A2 activities of Echis carinatus venom was determined. We found that 1.27mg of IgY (generated against FCA coated venom) and 1.33mg of IgY (generated against Bentonite coated venom) was able to completely neutralize the lethal activity (2LD50) of Echis carinatus venom. Both chicken egg yolk antibodies effectively inhibited procoagulant activity and partially inhibited the Edema forming activity. IgY generated using Adjuvant (FCA) coated venoms showed very high anti-phospholipase A2 activity. In general, the results demonstrated the effectiveness of chicken egg yolk antibodies raised against adjuvant and bentonite coated venoms in neutralizing various pharamacological effects of Echis carinatusvenom.Keywords: Venoms, FCA, Bentonite, Chicken antibodies (IgY), PLA2 Introduction Snake envenoming is a major public health issue in the rural tropics with large numbers of envenoming and deaths. There are nearly 3000 different species of snakes found in the world of which approximately 300 are venomous. In India there are about 216 different species are found, of which 53 species are reported to be poisonous. The common poisonous snakes found in India are Cobra, Krait, Russell’s viper and Saw Scaled Viper (Bawaskar, 2004). About 35,000 to 50,000 people die of snakebite every year in India. Antivenom immunotherapy is the only specific treatment against snake venom envenomation. In India polyvalent antisnake venom effective against venoms of Cobras, Krait, Russell's viper and Saw-scaled viper is available. Each ml of polyvalant Antisnake venom can neutralise 0.6mg of Cobra, 0.6mg of Russell’s viper, 0.45mg of Krait and 0.45mg of Saw Scaled Viper venom. Commercially available Horse antivenom contained high concentrations of non-immunoglobulins which frequently caused complement mediated side effects, serum sickness and renal failure which may be reduced by using sufficiently pure antibodies (Mayadevi et al., 2002). Thalley and Carroll (1990) described a new avian source of antivenoms that precludes these complications and an efficient method for preparing antivenoms composed solely of venom specific antibodies. Almeida 1998 reported that adult white leghorn hens hyperimmunized with Brazilian snake venoms produced antibodies capable of recognizing, combining with and neutralizing the toxic and lethal components of the venoms. The present study involves the comparison of the effectiveness of antivenom antibodies generated in chicken immunized with Freund’s complete adjuvant and bentonite coated venoms of Echis carinatus venom and the ability of these antibodies in neutralizing the various pharmacological activities induced by Echis carinatus venom was carried out by both in vivo and in vitro methods. Materials and Methods Venom and Experimental animalsThe free-dried snake venom powder of Echis carinatus venom was obtained from Irula’s Snake Catchers Industrial Co-operative Society Limited, Chennai and was stored at 4oC. Twenty four week old, single comb white leghorn chickens obtained from the Abinaya Poultry farm, Namakkal were maintained in our animal facility. They were used in the study for the production of antivenom (IgY). Male inbreed Swiss albino mice 18-20 gm were used for the studies of venom toxicity and in the experiments of venom neutralization. Institutional Animal Ethics Committee clearance at Institute of vector control and Zoonooses, Hosur, was obtained to conduct the experiment. All the animals were conditioned in standard cages.Development of antivenom antibodies in chick