Alum adjuvant is more effective than MF59 at prompting early germinal center formation in response to peptide-protein conjugates and enhancing efficacy of a vaccine against opioid use disorders

Figures & data

Figure 1. Behavioral, pharmacokinetic, and B cell comparison of alum and MF59. BALB/c mice were immunized with oxycodone vaccine OXY-KLH plus alum, OXY-KLH plus MF59, or a KLH control. (a) Mice were immunized on days 0, 14, 28, and 56, and OXY-specific IgG titers at 59 days were measured by ELISA (n = 7 per group). (b-c) Mice were immunized on days 0, 14 and 28, and motor activity induced by 5 mg/kg oxycodone was recorded over 60 minutes post-injection on day 45 (n = 7 per group). (c) Cumulative motor activity as total distance travelled during the first 15 minutes post-injection. (d) Mice were immunized on days 0, 14 and 28 and challenged with 2.25 mg/kg oxycodone on day 35, and oxycodone concentrations in brain 30 minutes after oxycodone administration were measured by GC-MS. (e) Mice were immunized on days 0, 14 and 28, and lymph nodes and spleens were collected at 14 or 42 days post-immunization (n = 5 per group). Total OXY-specific B cells populations were analyzed by flow cytometry. Data are mean±SEM. *p < 0.05, **p < 0.001, **** p < 0.0001.

Figure 2. Long-term flow cytometry analysis of 2W1S:I-Ab-specific CD4⁺ T cells using a peptide vaccine formulated with alum or MF59. Model immunogen phycoerythryin (PE) conjugated to streptavidin (SA), then conjugated to a model T cell peptide (2W1S) was used to track 2W1S:I-Ab-specific CD4⁺ T cells post-immunization. C57Bl/6 mice were immunized at days 0, 14 and 28, and cells were harvested at time points between 7 to 100 days (n = 3–6 per time point). (a) Total 2W1S:I-Ab-specific CD4⁺ T cells in the lymph nodes and spleen between 7 and 100 days post-immunization. (b) T follicular helper (Tfh) cells as percentage of 2W1S:I-Ab-specific CD4⁺ T cells. (c) GC-Tfh cells as a percent of 2W1S:I-Ab-specific CD4⁺ T cells. (d) Distribution of T cell subsets within the 2W1s:I-Ab-specific CD4+ T cell subsets at 7, 14, 42 and 100 days. Data are mean±SEM. *p < 0.05.

Figure 3. Flow cytometry analysis of OXY-specific B cells and 2W1S:I-Ab-specific CD4⁺ T cells in elderly mice. Elderly C57Bl/6 mice (aged 9–10 months) were immunized with OXY-KLH or ^2W1SSA-PE, and lymph nodes and spleens were collected and analyzed at 7 days post-immunization. (a) Proportion of OXY-specific B cells subsets in mice immunized with OXY-KLH formulated with alum or MF59 (n = 5 per group). (b) Proportion of T cell subsets in mice immunized with ^2W1SSA-PE formulated with alum or MF59 (n = 5 per group).