A step-by-step procedure is described for functionalizing the surface of a glass coverslip so that a single cell contacts distinct patterns of extracellular matrix and cellcell adhesion proteins. This dual-micropatterned substratum is accomplished through a two-step process. First, extracellular matrix (ECM) is microcontact-printed onto a silanized glass surface using electron beam lithography, etched resist-coated wafers, and Polydimethylsiloxane (PDMS) stamps of differing geometries. Then, non-ECM-coated surfaces are incubated sequentially with biotin, NeutrAvidin, and biotinylated Protein A to attach Fc-cadherin fusion proteins, Fc, or PEG. Cells are seeded at low density onto the functionalized surface for single-cell analysis of protein recruitment/turnover and cellular motility.
Lowndes, Molly, W. James Nelson
Adhesion Protein Protocols