Web Release Date: January 17,
Use of Photolithography to Encode Cell Adhesive Domains into Protein Microarrays
and
Departments of Biomedical Engineering and Chemistry, University of California, Davis, Davis, California 95616
Received September 17, 2007
In Final Form: November 5, 2007
Abstract:
Protein microarrays are rapidly emerging as valuable tools in creating combinatorial cell culture systems where
inducers of cellular differentiation can be identified in a rapid and multiplexed fashion. In the present study, protein
microarraying was combined with photoresist lithography to enable printing of extracellular matrix (ECM) protein
arrays while precisely controlling "on-the-spot" cell-cell interactions. In this surface engineering approach, the
micropatterned photoresist layer formed on a glass substrate served as a temporary stencil during the microarray
printing, defining the micrometer-scale dimensions and the geometry of the cell-adhesion domains within the printed
protein spots. After removal of the photoresist, the glass substrates contained micrometer-scale cell-adhesive regions
that were encoded within 300 or 500 
m diameter protein domains. Fluorescence microscopy and atomic force
microscopy (AFM) were employed to characterize protein micropatterns. When incubated with micropatterned surfaces,
hepatic (HepG2) cells attached on 300 or 500 m diameter protein spots; however, the extent of cell-cell contacts
within each spot varied in accordance with dimensions of the photoresist stencil, from single cells attaching on 30
m diameter features to multicell clusters residing on 100 or 200 m diameter regions. Importantly, the photoresist
removal process was shown to have no detrimental effects on the ability of several ECM proteins (collagens I, II,
and IV and laminin) to support functional hepatic cultures. The micropatterning approach described here allows for
a small cell population seeded onto a single cell culture substrate to be exposed to multiple scenarios of cell-cell
and cell-surface interactions in parallel. This technology will be particularly useful for high-throughput screening
of biological stimuli required for tissue specification of stem cells or for maintenance of differentiated phenotype
in scarce primary cells.
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