Assessment of the anti-Naja haje antibodies elicited in a low dose multi-site immunization protocol.

The horse antibodies to Naja haje (Nh) elicited in a low dose multi-site immunization protocol were investigated from binding perspective in the context of antivenom maturation. We found that, this protocol evoked the production of lethality neutralizing avid antibodies in the first round of immunization which increases over the successive immunization rounds. The changes and the relative changes in the antibody parameters of each horse were taken as a measure for the efficacy of its immune system to respond by gaining and magnifying the antivenom parameters. While the avidity increases over the successive immunization rounds towards a maturation ceiling, the avidity gain and relative gain decreases. While the ED(50) gain increases over the immunization rounds, the relative ED(50) gain of the strong responder horse-1 is steadier than for the weak responders horse-2 and horse-3. The profiles for the gain and the relative gain of ELISA titers by three horses are pronouncedly different, reflecting great variations in the immune response by the individual animals. A late antigen dependent avidity pulse was detected after the last dose of the second immunization period, demonstrating that antibody maturation continues during the rest periods. The avidity pulse appears as an increase in avidity which returns to a base level within four weeks. These results indicted that maturation of antibodies to lethality neutralization relevant venom epitopes rather than maturation of antivenoms is the most important potency determining factor. Eventually, a main advantage of the low dose multi-site immunization protocol is the production of highly avid antibodies at the primary immunizations which is mostly crucial for the production of highly potent antivenoms.