Fig. 3
Antivenomics analyses of crotalic antivenom manufactured by IVB from optimized venom mixture. a and d about 300 μg of crotamine-positive and crotamine-negative venoms separated by RP-HPLC, respectively. b and e the RP-HPLC profiles of immunocaptured fractions of crotamine-positive and crotamine-negative venom from affinity column with crotalic antivenom SAC155204F (raised against a pool of both types of venoms), respectively. c and f the flow-through fraction of crotamine-positive and crotamine-negative venoms, respectively. The elution conditions were: isocratic gradient with 5% of 0.1% TFA in 100% of acetonitrile (solvent B) for 5 min, followed by 5-25% B for 5 min, 25–45% B for 60 min, 45–70% B for 10 min, with a final isocratic step of 70% B for 5 min at a flow rate of 1 mL/min. The first peak present in all chromatograms (elution time ~ 3 min) is a technical artifact. Protein families associated with HPLC peaks: 1 – crotamine, 2 – disintegrin, 3 – crotoxin acid chain 4 to 7 – crotoxin basic chain and 8 – fractions of low expression toxins including D49-PLA2, serine protease (gyroxin), C-type lectin (convulxin) and PIII-metalloproteases