Such efforts have paid off, with 180,912 children immunized by the second immunization round in September – amounting to 102 per cent of the original target.
The HA1-2-specific IgG titers induced by HA1-2-fliC after the third immunization were about 4-fold higher than those following the second immunization (Fig. 6a).
ELISpot assays were conducted using fresh PBMCs collected pre-immunization (day 0), 2 weeks after the second immunization and 2 weeks after the third immunization, by stimulating with individual peptides.
One week after the third immunization, serum antibody titers of immunized mice were determined by indirect ELISA on inactivated E. tarda or OMP-coated plates.
Solicited adverse events were recorded by direct questioning on days 0, 2, 7, 10, 14 and 28 following the first and second immunizations and on days 0, 2, 7, 10 and 14 following the third immunization, and by a diary card filled out days 1 through 7 after each immunization to record oral temperature.
However, unexpectedly severe responses after priming forced us to delay the second immunization by three weeks and ultimately to eliminate the third immunization.
This cross-reactivity, which was not related to a known history of malaria exposure, was exceptionally boosted already by the first immunization, yielding the highest titers observed in the entire trial.
Blood was collected 14 days after the second immunization and 7 days after the third immunization from all groups for analysis by microneutralization assay and ELISA.
After the third immunization, functionality of the antibody was tested by an in vitro growth inhibition assay.
