Nanoscale characterization of cell receptors and binding sites on cell-derived extracellular matrices

Cells are able to adapt their extracellular matrix (ECM) in response to external influences. For instance polymer scaffolds with tunable properties allow for guiding cell adhesion behavior and ECM adaptation in a controlled manner. We propose a new and versatile approach for the investigation of extracellular molecular assemblies at materials interfaces by scanning force microscopy. The distribution of cell adhesion receptors and binding sites of matrix proteins in the investigated ECMs was identified by immunolabeling with 15 nm gold beads. To precisely localize the immunogold in the matrices we utilized electrostatic force microscopy that allows for materials-dependent contrast according to differences in the dielectric properties of the immunolabels. In addition, an image processing routine was developed to localize the immunogold by correlation analysis. The applicability of our approach for nanoscale characterization of cell-derived ECM was further verified in two independent experiments. We probed the distribution of the cell adhesion receptor α(5)β(1) integrin next to its extracellular ligand fibronectin and the corresponding binding site on the fibronectin molecule.