Figure 3.
Integrin forces unfold I27 through RGD and FN. (A) Illustration of the RGD-Cy3-I27 MTFM sensor containing the p-azidophenylalanine amino acid. The dye was conjugated with Cu-free cycloaddition and was subsequently immobilized onto the AuNP surface. The quenching efficiency was 92% (top). Representative RICM, GFP-paxillin, Cy3 tension and overlay of Cy3/GFP shows colocalization of FAs with tension. (B) FN (9–10) sensor was labeled with the Alexa 647 and co-incubated on the AuNP surface with RGD-I27-Cy3 for 1 hr (FN 9–10 PDB ID: 1FNF). Representative RICM and fluorescence images of a REF cell cultured on a binary tension probe surface for 1 hr. The RGD-Cy3-I27 and FN-A647-I27 fluorescence signals are displayed from the highlighted region of interest. Overlay of these channels and line scan analysis shows colocalization of both sensors. Scale bar, 10 μm.