Figure 5. TCR tension at T-B cell interfaces.

a Schematic showing the functionalization of B-cell membrane with DOTS. Locking strand was used to capture the tension signal at the T-B cell interface. b-c Representative images showing the DOTS and tension signals at the T-B cell interfaces. DOTS were modified with anti-CD3ε (b) or pMHC (c). d-e Plot comparing the Atto647N/Cy3B ratios on the B-cell surface and T-B interfaces with and without locking strand after 30 min incubation. B-cells are modified with anti-CD3ε DOTS (d) or pMHC DOTS (e). Data from each group was acquired >25 T-B cells. f Schematic showing that TCR-anti-CD3ε has a higher bond lifetime than TCR-pMHC and thus scans and lights up fewer DOTS within a specific period (30min). g Flow cytometry fluorescence plots and forward scatter plots were used to gate out the B-cell monomer and T-B 1:1 binding population. h-i Atto647N fluorescence histograms of B-cells (blue) and T-B 1:1 conjugates (red) with and without locking strand. Plot quantifying Atto647N MFI difference between B-cells and T-B cell 1:1 conjugates with and without locking strand present in the media. Data were acquired from 4 independent experiments and 4 independent mice. B-cells are modified with anti-CD3ε DOTS (h) or pMHC DOTS (i). Scale bars = 5 μm.