Management of Snakebite and Research Management of Snakebite ...

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MATERIALS AND METHODS Method (1) Nitrocellulose paper 0.22µ (2) Russell’s viper venom (RVV) (Myanmar Pharmaceutical Factory) (3) Bovine Serum Aibunim fraction V (4) Affinity purified rabbit anti-RVV IgG (5) Dye antibody conjugate The principle of the test is double antibody sandwich dot blot. Preparation of a dipstick A nitrocellulose strip is coated with rabbit anti-RVV IgG antibody and blocked with 5%Bovine Serum Aibunim in Phosphate Buffered Saline. Then, it is incubated with venom standard (60ng/ml) and human serum (negative control) for 15 min. at room temperature and rinse with tap water and dry. It is stored in a sealed polythene bag at 4°C (six months) and room temp (three months). Testing of a patient’s serum sample A test strip is laid on a flat surface, add one drop of serum to the test spot and incubate for 15 min. at room temperature. Then, it is rinsed with tap water, dried and add one drop of dye antibody conjugate to test, standard and control. If it is positive, a red ring develops. Interpretation A red ring indicates a venom level of 10 ng/ml or more. The venom level is estimated by comparing the colour intensity of the ring with the standard (60ng/ml Russell’s viper venom). No colour develops in control. • An insulin syringe is used for delivery of sample and dye conjugate • Dye antibody conjugate is freshly reconstituted with 100 µl of the diluent, stored at 4°C and used within two weeks. RESULTS A. Sensitivity of the test –10ng/ml of RV venom B. Specificity–does not cross react with other snake venoms. A Dipstick Test for Rapid Diagnosis of Russell’s Viper 107
Management of Snakebite and Research 108 Performance of the test Colloidal Dye Immunoassay (Dipstick Test) DISCUSSION Enzyme Immunoassay Positive Negative Specificity 100% (95% CI 81.5-100) Sensitivity 97.0% (95% CI 88.7-99.5) Positive predictive value 100% (95% CI 93-100) Negative predictive value 91.7% (95% CI 71.5-98.5) Kappa coefficient 0.9414 One tailed p value 0.0000 (
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