OT-I–GFP T cells reduced crawling speed, increased frequency of stopping, and increased DC contact duration after antigen recognition. Representative video of OT-I–GFP Rag2 −/ − and polyclonal T cell interactions with DCs before and after antigen addition. CMTMR-labeled OT-I–GFP Rag2 −/ − cells (red) and CFP+ polyclonal CD8 T cells (blue) were cotransferred into CD11c-YFP (green) recipients, and recipients were immunized with CFA in the footpads. 18 h after T cell transfer and immunization, the draining popliteal LN was explanted and visualized by two-photon microscopy before and after the addition of SL8 peptide antigen. Imaging was performed in the most superficial 80 µm of the LN below the capsule to replicate imaging conditions during TCR visualization. The video represents a maximum intensity z projection of an imaging volume of 100 (x) × 100 (y) × 80 µm (z). Elapsed time is displayed in minutes:seconds..
