Figure 6.

Eye drop instillation of ovalbumin with cholera toxin adjuvant elicits a germinal center reaction in TALT and draining lymph nodes. (A) Immunization scheme of eye-drop immunization of BALB/c mice with PBS, OVA, or OVA with CT. (B) Confocal microscopic analysis of TALT sections stained with a lectin, PNA (green), and anti-B220 antibody (blue). Arrowheads show germinal center (PNA⁺ B220⁺ cells). Bars: 50 µm. (C) Expression of CD95 and GL7 on TALT cells pooled from the 4 or 5 mice. Numbers in outlined areas show the percent of germinal center (GC) B cells (CD95⁺ GL7⁺ cells) in CD3e^- B220⁺ lymphocytes in TALT. (D) The number of GC B cells and Tfh cells (CXCR5⁺ PD-1⁺ in CD4⁺ CD8^- T cell) in the cervical and submandibular lymph nodes (cLN and smLN) was quantified by flow cytometry at 1 week after the final immunization. Gating strategies for the flow cytometry analyses were shown in Supplementary Figure 7 . (E) OVA-specific serum IgG levels and tear IgA levels were measured by ELISA. Data were obtained from a single experiment. Bars represent the mean ± standard deviation. ***P < 0.001, *P < 0.05 (GC B cell, OVA-specific serum IgG and tear IgA: Kruskal-Wallis test, Tfh cell: one-way ANOVA by Tukey’s test, PBS: n = 4 or 5, OVA: n = 5, OVA + CT: n = 5 animals).