Class II MHC–independent contacts and suppression of DCs by T reg cells. (A) The protocol for visualizing T reg cells interact with MHC class II sufficient or deficient DCs in vivo. (B) Interactions between T reg cells and DCs deficient in MHC class II (I-A^b) expression (MHCII^−/−) or WT DCs (MHCII^+/+) visualized in the same draining LN (see Video 10) for the corresponding image sequence. Arrowheads trace two T reg cells comparably interacting with two DCs of the two genotypes, respectively, with closed arrowheads specifically denoting the beginning and end of an interaction incidence. Time code, hour:min:sec. Bar, 15 µm. (C) Duration of T reg cell contacts with DCs of indicated types (52 MHC class II^+/+ and 47 MHC class II^−/−) pooled from two experiments. (D) The protocol for visualizing interactions between OT-I T cells and OVA_257-264-presenting DCs that in LN of Foxp3-GFP mouse. (E and F) Typical behaviors of OT-I T cells interacting with DCs visibly contacted or untouched by the Foxp3⁺ T reg cells. Closed arrowheads trace three OT-I T cells making contacts with DCs. Two (cell #1 and #3) exhibit shorter contacts with a DC touched by a T reg cell (open arrowhead), and one (cell #2) was engaged in long-lasting contact with a DC not touched by any visible T reg cells during the session. (F) Duration of contacts between OT-I T cells and DCs with (n = 52) or without (n = 50) T reg cell contacts. Data are pooled from two independent experiments (see Video 11) for the corresponding image sequence. ***, P < 0.001; N.S., not significant, Student’s t test.