Figure 5.

Conjugate vaccines produced using the MD min CFE formulation elicit antibodies that are bactericidal. (A) Lyophilized MD min CFE reactions using iVAX extracts were used to synthesize anti-ETEC O78 conjugate vaccines for immunization studies. Groups of BALB/c mice were immunized subcutaneously with a 1:1 mixture of adjuvant and PBS or ∼24 μg of the following cell-free-derived immunogens: unconjugated protein D (PD), or PD modified with O78 O-PS from a minimal iVAX reaction (PD-O78 (MD min)). Each group was composed of eight mice. Mice were boosted on days 21 and 42 with identical doses of antigen. (B) ETEC O78 O-PS-specific IgG titers were measured by enzyme-linked immunosorbent assay (ELISA) in endpoint (day 56) serum of individual mice (black dots) with recombinant O-PS immobilized as antigen. Mean titers of each group are also shown (red lines). Statistical significance was calculated by unpaired two-tailed t-test with a single asterisk (*) indicating p-value <0.05 and ns indicating not significant. (C) Bacterial killing activity of serum antibodies corresponding to the same groups as in (B). Survival data were derived from a standard serum bactericidal assay (SBA) where dilutions of pooled sera from immunized mice were tested against ETEC O78 strain H10407 in the presence of human complement. Values for % survival were determined from the colony forming units (CFUs) counted at each individual serum dilution. Data in (C) represent average error bars for two independent samples (n = 2).