Management of Snakebite and Research Management of Snakebite ...

Conventional RVV toxoid
In 1969, Ko Ko Gyi et al tried to prepare RVV toxoid from desiccated RVV in the veterinary
Department of BPI (now known as MPF), Yangon. RVV was dissolved in one percent
phosphate buffer saline solution (pH 7.2) containing one percent peptone, to give 1 mg/
ml concentration. Formalin was added to give 0.5 percent concentration and incubated at
37° C for four weeks. The toxicity of the detoxified venom was reduced to 60 times and
100 times over that of the initial venom when tested with rabbit and mouse respectively.
Beside, rabbits inoculated intraperitonally with 5 ml of detoxified venom at three days’
interval for four times were found to be able to withstand 2-3 times the minimal lethal
dose of venom when challenged seven days after the last immunizing dose. The results
presented herein demonstrate that it is possible to detoxify RVV with formalin without
effecting its immunogenicity. However, further detailed investigations of the toxoid regarding
its stability, the level and persistence of circulating antibody, chronic toxicity of the toxoid
on vital organs following immunization could not be carried out.
Refined RVV toxoid
Development of Russell’s Viper Toxoid
In 1986, the Department of Medical Research (DMR) working group on RVV toxoid,
successfully developed a refined RVV toxoid. The desiccated pooled crude RVV was
fractionated using sephadex G50 fine with 0.01mol/l Tris-HCL buffer pH 7.5. The first four
fractions obtained were pooled and concentrated and a mild and step wise formalin
toxoiding was carried out by using a modified method of Kondo et al (1971).
This preparation was found to be safe, since mice tolerated 0.5ml of toxoid (i.e. more
than 60 LD50. The antibody titers of mice and rabbits, which were immunized with toxoid
subcutaneously for three booster injections at weekly interval commencing three weeks
after the first injection, showed a rise of 1.2 gm % to 4 gm % in their sera, as shown by
quantitative reversed rocket immunoelectrophoresis method. The immunized rabbit could
withstand the challenge of 8 mg to 16 mg of RVV by intramuscular injection. The antibody
titer estimated by the in vivo venom neutralization test revealed that 0.4 ml serum neutralized
200 to 300 mg 0f venom.
Monkeys (Macaca mulata) were immunized with an initial subcutaneous injection of
0.5 ml (containing 1 mg protein) by two boosters of same dosage given at four week
intervals. Satisfactory rise of circulating antibody levels was seen in these monkeys and
their rise persisted for 24 weeks. The immunized monkeys withstood intramuscular
challenged with 2 mg/kg of crude venom (i.e. lethal dose) at two weeks after a course of
toxoid injection. It also showed minimum undesirable local side effect with no detectable
systemic adverse effect.
In another experiment, refined toxoid was given to 12 human volunteers. No
abnormalities such as bleeding time, clotting time, haemoglobin concentration, total
differential white cell count, serum bilirubin, alkaline phosphates, alanine and aspartate
transferase, urinalysis and ECG done before injection and 2, 7 and 21 days after injection
of refine toxoid were detected in the laboratory test. Significant rise of antibody titer was
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