| Abstract |
Here we describe a protocol to prepare cell patterns by immobilizing anchor-dependent cells onto the desired area of an unpatterned culture substrate by micro-patterned light projection with high precision to the size of one single cell. The method is based on the phenomenon that the Ca(2+)-dependent adhesion of anchor-dependent cells becomes insensitive temporarily to the removal of Ca(2+) due to the influence of the less-invasive in situ UV irradiation. By repeating the process, a patterned coculture system can be fabricated. In clear contrast to conventional cell patterning that uses fixed-patterned substrates, the cell-retaining area is defined after cell seeding. Therefore, the cell separation on a substrate based on imaging cytometry can be implemented by combining this method with a computer-controlled microprojection coupled with a microscope. Because the cells thus patterned can continue to grow and migrate freely afterward from their initial positions, the method is expected to provide a useful cell manipulating tool, especially in cases where spontaneous development of the cells after patterning is necessary and important, such as an analysis of cell migration.
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