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. 2011 Jan 17;6(1):e14498. doi: 10.1371/journal.pone.0014498

Figure 2. Concentration of RTX at RAW-Ramos interfaces.

Figure 2

(A) Volumetric reconstruction from confocal slices of a Ramos-RAW cell interface. RTX-Al488-coated (green), PKH26-labelled Ramos cells (red) were incubated with RAW cells for 45 minutes at 37°C. RAW cells were labelled with anti-CD11b-APC (cyan). The RAW cell has extensively trogocytosed both RTX and PKH26. Inset shows the dotted area above it without the PKH26 channel overlaid, revealing the concentration of RTX-Al488 at the cell-cell interface, otherwise depleted from the rest of the Ramos cell. Trogocytosis reaction was halted by fixation 45 min after co-incubation. Ramos cells are approximately 12 µm in diameter. (B) Deconvolved epifluorescence images from live experiment of PKH26-labelled, RTX-Al488 coated Ramos after coincubation with RAW cells for 1 hour. Extended focus and volumetric representations of the imaged slices show accumulation of RTX-Al488 at the interface. Scale bar 5 µm. Unit size for volume 3.3 µm. (C) Frames from live microscopic imaging following a RTX-Al488 coated Ramos cell settling onto a RAW macrophage. Extensive RAW membrane ruffling occurs to capture the uropod (black arrow, 9 minutes), and RTX is internalized shortly after (white arrow, 13.5 minutes). See Videos S3. Scale bar 10 µm.