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Figure 4


Fig. 4. Single-cell force spectroscopy (SCFS) to study the adhesive strength of EphBR–ephrin-B2 interactions in a cellular context. (A) Model of SCFS. An individual J774 cell was attached to a cantilever and moved towards the surface of HUVECs that overexpressed ephrin-B2 or {Delta}C-ephrin-B2, or towards mock-transduced HUVECs. The cantilever was then moved in the opposite direction to separate the cells following a cell-cell contact time of 1 minute. The adhesive force was calculated from the deflection of the cantilever and its spring constant. (B) J774 cell attached to the tip of the cantilever (left). Cantilever above the HUVEC layer (right). (C) Average maximum unbinding force and work needed to detach the J774 cell from ephrin-B2- or {Delta}C-ephrin-B2-transduced HIVECs or from mock-transduced HUVECs. Statistical analysis using a one-way ANOVA followed by a multiple pairwise comparison (Holm-Sidak) revealed that there was no significant difference between either ephrin-B2 or {Delta}C-ephrin-B2 whereas the difference between mock and both experimental groups was significant (*P<0.05). Average values were calculated from n=24-30 individual force-distance measurements. The average work required to separate the cells completely from each other was represented by the area under curve of the force plot. Scale bars: 50 µm.





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