Traditional, horse-derived
antivenin is currently the most efficient treatment against
snake bites. However, it is costly and has unpredictable side effects. Thus, alternative, cost-effective strategies for producing
antivenin are needed. In this study, we immunized hens with inactivated NNA
venom proteins from the cobra Naja naja atra (NNA). Purified yolk
IgY antibodies showed specific anti-NNA binding activity comparable to that of the equine-derived
antivenin. We used phage display technology to generate two antibody libraries containing 9.0 × 10⁸ and 8.4 × 10⁸ clones with a short or long linker, respectively. The phage ELISA indicated that anti-NNA clones displaying
single-chain variable fragments (scFv) were significantly enriched after biopanning. The nucleotide sequences of the light and heavy chain genes of 30 monoclonal
scFv antibodies were determined and classified into six groups with the short linker and nine groups with the long linker. These scFv clones specifically bound to NNA
proteins but not to
venom proteins from other snakes. Their binding affinities were further determined by competitive ELISA. Animal model studies showed that anti-NNA
IgY antibodies exhibited complete protective effects, while a combination of
scFv antibodies raised the survival rates and times of mice challenged with lethal doses of NNA
venom proteins.