CXCR3-dependent clustering of viral-specific CD4⁺ T cells in mIFRs. (a) Flow cytometry of OT-II T cell activation after UV-PR8-OTII vaccination at the indicated time points (n = 4). (b) Expression of CD69 (black) and CXCR3 (red) by OT-II T cells at the indicated time points as in panel a. CD69⁺ OT-II cells were gated for CXCR3 expression analysis. Symbols represent individual mice. ANOVA (CD69), P < 0.0005; ANOVA (CXCR3), P < 0.005 (n = 4). Mean ± SEM. (c) CXCL10 expression in vaccinated REX3 mice or unvaccinated REX3 controls by flow cytometry. (left) Displayed cells gated on CD11c^hi cDCs, followed by CD11b^hi or CD8a⁺ as indicated. (right) Percentage of CXCL10-positive DCs by subset after vaccination (n = 4). ***, P < 0.001. Horizontal bars indicate mean. (d) MP imaging of REX3 PLN 24 h after UV-PR8 vaccination. Dashed line: medulla (M). B, follicles; T, T cell cortex. Images are representative of four PLNs from two independent trials. (e) Confocal imaging of REX3 PLN 24 h after UV-PR8 vaccination. Arrowheads: CXCL10-positive DCs. Images are representative of four PLNs; two independent trials. (f) C57BL/6 recipients received differentially labeled WT or CXCR3-deficient naive OT-II T cells. Recipients were vaccinated with UV-PR8-OTII, and PLNs were collected at 24 h. Quantitation of clustering efficiency of WT or CXCR3-deficient OT-II T cells is shown (n = 5). (g and h) Adoptive transfers were performed as in f. Recipients were vaccinated, and PLN suspensions were collected 60 h after vaccination for flow analysis. Individual T cell populations were compared with the same population in unvaccinated contralateral controls. CD40L (g) and CD69 (h) acquisition is expressed as fold change (FC) in vaccinated versus unvaccinated population controls (n = 5). **, P < 0.005; ***, P < 0.001.