Figure 3: Quantitative analysis of integrin at actin nodules using dSTORM.

(a) Human platelet stained with Alexa488-phalloidin and imaged for F-actin using diffraction limited TIRF to identify the nodules (left panel) and Alexa647-anti-αIIb and imaged using super-resolution dSTORM TIRF (right panel). Actin nodules and the corresponding integrin depleted zones are indicated by arrows. (b) Quantitative cluster mapping analysis of integrin localization on the platelet surface (different cells to that shown in part (a)). One micrometre square regions of interest were taken around regions without (top row) and with (middle and bottom rows) actin nodules. For each sample, the left hand panel shows the dSTORM picture, the middle panel shows the cluster heat map and the right hand panel shows the thresholded image showing the integrin depleted zones detected in the distribution of integrin on the platelet surface. The dotted yellow line indicates the position of the F-actin dense core. The actin nodule in the middle row shows no clustering of the integrin around the nodule whereas the one seen in the bottom row appears to show integrin clustering (arrows). Scale bar, a: 2 μm, b: 0.5 μm.