Fig. 4. Molecular composition of focal contacts on micropatterned substrata. B16 cells were cultured for 1 hour on patterned ECM substrata and labelled for focal adhesion molecules and ECM proteins. (A,A') Cell on a homogenous fibronectin substratum prepared with µCP. Fluorescence staining for vinculin (Vin, green) and actin (Act, red) reveals a staining pattern of dot-like or elongated adhesion foci mainly at the cell periphery that were connected to actin bundles. (B,B') Cell stained for vinculin (Vin, green) and actin (red) on a patterned substratum of 0.6 µm² fibronectin dots (FN, blue). Vinculin has accumulated in areas of the cell overlying ECM dots. Actin fibres terminate in most of these adhesion sites, indicating functional contact sites. (C,C') Cell stained for focal adhesion kinase (FAK, green) on a patterned substratum of 1 µm² FN dots (red). (D,D') Cell stained for phosphotyrosine (PT, green) on a patterned substratum of 1 µm² FN dots (red). (E,E',E'') B16 cell expressing ß3-integrin-GFP (green) stained for paxillin (Pax, red) on a patterned vitronectin (VN, blue) substratum of 0.6 µm² dots. (F,F') B16 cell expressing ß 3-integrin-GFP (green) labelled for actin (red) growing on vitronectin (blue) at the border between a uniform and a patterned substratum of 1 µm² dots. Note the redistribution of integrin receptors on the patterned substratum. Scale bars: 10 µm.