Fig. 4.

Local Ag presentation during the effector phase is required for DLN and spleen T_FH as shown by Ag delivery via i.n. vs. i.s vs. i.v. routes. (A) Experimental design: OVA_II peptide-pulsed B cells (CD45.1⁺ or GFP⁺) were used as APCs and transferred into PR8 infection-matched hosts 6 dpi by either i.n., i.s., or i.v. routes. Unpulsed APCs were transferred both i.n. and i.s., or both i.n. and i.v., as negative controls. In vivo-generated 6-d OT-II.Nur77^GFP.Thy1.1⁺ effectors were transferred by i.v. route. Mice were harvested 14 to 16 h posttransfer (pt), and donor cells were analyzed by flow cytometry. (B and H) Number of transferred APCs. (C and G) Donor cell Nur77^GFP expression was analyzed by flow cytometry in the lung, DLN, and spleen. (n = 8 to 11 per group pooled, 3 independent experiments). (D–F, I, and J) Experiment was performed as in A, and mice were euthanized 3 to 4 dpt. (D and I) Total numbers of DLN and spleen donor effectors recovered (n = 5 to 12 per group pooled, 3 to 4 independent experiments). (E and J) DLN T_FH formation from donor cells (n = 5 to 11 per group pooled, 2 to 3 independent experiments) (F and K) Spleen T_FH formation from donor cells (n = 5 to 12 per group pooled, 2 to 4 independent experiments).