Evaluation of a nanostructured CpG-ODN/ascorbyl palmitate as a safe and effective adjuvant for anticrotalic PLA2 serum.

Background: The WHO states that antivenom is the only safe and effective treatment to neutralize snake venom. Snakebite antivenom typically involves horse hyperimmunization with crude venom and Freund's adjuvant.

Methods: In the current work, we analyzed the ascorbyl palmitate liquid crystal structure with snake protein or PLA2, the carrier charge capacity, and we evaluated the immune response induced by the enzyme P9a(Cdt-PLA2) formulated in a nanostructure using CpG-ODN, determining the titer of IgG antibodies. BALB/c mice were subcutaneously immunized on days 0, 15 and 30 with P9a(Cdt-PLA2)/CpG-ODN/Coa-ASC16 or P9a(Cdt-PLA2)/Freund's adjuvant (complete first and incomplete-booster). On day 48 the mice were sacrificed. The neutralization ability of antibodies from animals immunized with P9a(Cdt-PLA2)/CpG-ODN/Coa-ASC16 or P9a(Cdt-PLA2)/Freund's adjuvant was tested against PLA2 activity and venom lethality.

Results: In both groups of immunized mice, the antibody titers in blood samples at the assayed time were high (approximately 1×105). The antibodies were able to neutralize P9a(Cdt-PLA2) activity in vitro and lethality in vivo. Microscopic analysis showed that P9a(Cdt-PLA2)/CpG-ODN/Coa-ASC16 produces minimal damage at injection sites compared with Freund's adjuvant.

Conclusion: The Coa-ASC16/CpG-ODN formulation shows promise as a safe and effective adjuvant against crotalic PLA2, inducing a strong humoral response and reducing local tissue damage compared with Freund's adjuvant.