Figure 1. Bisphosphonates Increase Antibody Responses to Live and Inactive Viruses, Proteins, Haptens and Existing Commercial Vaccine Formulations.

(a) VSV nAb titers in the serum of C57BL/6 mice that were footpad injected with PBS, PBS-liposomes (PBS-Lip), clodronate (CLD) or clodronate liposomes (CLD-Lip) prior to VSV infection in the same footpad (see Methods for details). n = 5 per group. Black asterisks, PBS versus CLD-Lip; red asterisks PBS versus CLD. Results are representative of >10 independent experiments. (b) VSV nAb titers 7 days p.i. in the serum of C57BL/6 mice that were footpad injected with CLD, etidronate (ETD), pamidronate (PMD) or alendronate (ALD) prior to VSV infection in the same footpad. n = 5 per group; results are representative of 3 independent experiments. (c) VSV nAb titers at day 14 post immunization in the serum of C57BL/6 mice that were footpad injected with PBS or CLD prior to paraformaldehyde-inactivated VSV (PFA-VSV) immunization in the same footpad. n = 10 per group; results are representative of 3 independent experiments. (d) Ovalbumin (OVA)-specific IgG titers at day 14 post immunization in the serum of C57BL/6 mice that were footpad injected with PBS or CLD prior to OVA immunization in the same footpad. n = 4 per group; results are representative of 3 independent experiments. (e) 4-Hydroxy-3-nitrophenylacetyl (NP)-specific IgG titers at day 14 post immunization in the serum of C57BL/6 mice that were footpad injected with PBS or CLD prior to NP-chicken gamma globulin (CGG) immunization in the same footpad. n = 5 per group; results are representative of 2 independent experiments. (f) HBsAb titers (mIU/ml) 21 days p.i. in the serum of C57BL/6 mice that were injected in the footpad with PBS or CLD prior to Engerix-B (an approved vaccine against hepatitis B virus) immunization in the same footpad. n = 5 per group; results are representative of 5 independent experiments. (g) H₁N₁-specific IgG titers in the serum of C57BL/6 mice that were injected intramuscularly with PBS or CLD prior to intramuscular immunization with H₁N₁. n = 5 per group; results are representative of 2 independent experiments. (h) Kaplan-Meier survival curves of C57BL/6 mice injected in the right footpad with PBS, 10⁶ pfu equivalents of PFA-inactivated VSV (PFA-VSV) or 10⁶ pfu equivalents of PFA-inactivated VSV and clodronate (PFA-VSV+CLD) 3 weeks prior to infection in the left footpad with 3.5 × 10⁸ pfu of VSV. n = 12 per group; PFA-VSV+CLD versus PFA-VSV: P < 0.05, Log-rank (Mantel-Cox) test. Results are representative of 2 independent experiments. (i) VSV nAb titers 125 days p.i. in the serum of C57BL/6 mice that were footpad injected with PBS or CLD prior to VSV infection in the same footpad. n = 5 per group; results are representative of 2 independent experiments. ***: P < 0.001; **: P < 0.01; *: P < 0.05 See also Figure S1.