Impaired primary, secondary, and memory responses to NP-OVA in P2rx7^−/− mice. (A) Scheme of the immunization protocol and time points of analysis. (B) Frequency and absolute number of CXCR5⁺ICOS⁺ Tfh cells in the draining lymph nodes from untreated or immunized WT and P2rx7^−/− mice at 1 wk after s.c. injection of NP-OVA in MF59 (WT unimmunized, n = 5; P2rx7^−/− unimmunized, n = 5; WT NP-OVA, n = 8; P2rx7^−/− NP-OVA, n = 8), representative contour plots of staining with human CLIP and OVA tetramers, and statistics of OVA tetramer positive cells (n = 4). (C) Frequency and absolute number of GC B cells (same samples as above) gated as PNA^hiB220⁺ among CD19⁺ B cells (upper bar graphs) and of NP-positive GC B cells (lower bar graphs) with representative contour plots. (D) Frequency and absolute number of NP-positive GC B cells in the spleen of the indicated mice within secondary response to NP-OVA. WT unimmunized (n = 2), P2rx7^−/− unimmunized (n = 2), WT NP-OVA (n = 4), and P2rx7^−/− NP-OVA (n = 4). Absolute number of NP-specific IgG secreting cells in the spleen (WT NP-OVA, n = 8; and P2rx7^−/− NP-OVA, n = 9) and BM (WT NP-OVA, n = 7; and P2rx7^−/− NP-OVA, n = 8) of immunized mice. (E) Frequency of OVA tetramer positive cells among CD4⁺ T cells (n = 5) and absolute number of NP-specific IgG secreting cells in the spleen of the indicated mice. WT and P2rx7^−/− no recall (n = 4), WT and P2rx7^−/− NP-OVA (n = 5). Unpaired Student’s t test. Each dot in graphs represents an individual mouse, and horizontal lines represent median values. *, P < 0.05; **, P < 0.01. ns, not significant.