Micropatterning of proteins and mammalian cells on indium tin oxide

This paper describes a novel surface engineering approach that combines oxygen plasma treatment and electrochemical activation to create micropatterned cocultures on indium tin oxide (ITO) substrates. In this approach, photoresist was patterned onto an ITO substrate modified with poly(ethylene) glycol (PEG) silane. The photoresist served as a stencil during exposure of the surface to oxygen plasma. Upon incubation with collagen (I) solution and removal of the photoresist, the ITO substrate contained collagen regions surrounded by nonfouling PEG silane. Chemical analysis carried out with time-of-flight secondary ion mass spectrometry (ToF-SIMS) at different stages in micropatterned construction verified removal of PEG-silane during oxygen plasma and presence of collagen and PEG molecules on the same surface. Imaging ellipsometry and atomic force microscopy (AFM) were employed to further investigate micropatterned ITO surfaces. Biological application of this micropatterning strategy was demonstrated through selective attachment of mammalian cells on the ITO substrate. Importantly, after seeding the first cell type, the ITO surfaces could be activated by applying negative voltage (-1.4 V vs Ag/AgCl). This resulted in removal of nonfouling PEG layer and allowed to attach another cell type onto the same surface and to create micropatterned cocultures. Micropatterned cocultures of primary hepatocytes and fibroblasts created by this strategy remained functional after 9 days as verified by analysis of hepatic albumin. The novel surface engineering strategy described here may be used to pattern multiple cell types on an optically transparent and conductive substrate and is envisioned to have applications in tissue engineering and biosensing.