Fig. 1. Intranasal immunization adjuvanted with S100A4 provokes greater humoral and cellular immune responses.
Mice were immunized three times intranasally (i.n.) with ovalbumin (OVA; 10 μg) alone, or admixed to S100A4 (10 μg) or cholera toxin (CT; 1 μg) at a 10-day interval. Unmanipulated naïve mice were included for baseline control. Mice were euthanized 10 days or 196 days after the last immunization for collection of various types of samples. a The immunization and tissue collection procedures are indicated using a flowchart. b–f Various classes of OVA-specific antibody levels in serum (b), lung exudates (c), broncho-alveolar lavage fluid (BALF) (d), vaginal lavage (e) and pooled feces (f) were analyzed by ELISA. g–i Single-cell suspensions were prepared from the lung tissue (g), lymph nodes (h), and spleen (i). Percentages of CD8 T cells that recognize H-2Kb-OVA (SIINFEKL) tetramer were revealed by flow cytometry. Contour plots indicating a representative mouse from each treatment (upper panel) and pooled data from all the mice (lower panel) are shown. Numbers adjacent to outlined areas indicate percent cells in each gate. Each dot represents data from an individual mouse and blue lines indicate the average values. #P = 0.0022 or the exact P values (italic numbers) are indicated by Mann–Whitney U test. Except for the long-term observation experiment, the other measurements represent at least three separate experiments.