Automated analysis and survival selection of anchorage‐dependent cells under normal growth conditions

Abstract

An instrument is described that can automatically analyze and select for a subpopulation of anchorage‐dependent cells in tissue culture. Cells that label with fluorescently tagged antibodies or demonstrate structural variations are saved from exposure to a destructive high‐intensity argon laser beam. The surviving population may then be cloned. The cell selection may occur in a tissue culture plate or in a microflow incubator which is designed to maintain a constant flow of media at 37°C across cells growing on a glass coverslip. This incubator sits on an inverted microscope which focuses the laser beam to a diameter as small as 1 μm. A high‐speed computer‐controlled two‐dimensional stage moves the cells past the beam for analysis, the results of which determine the fate of each cell: whether it is to be destroyed by radiant energy or selected for survival and subsequent proliferation. Another selection strategy performed by the instrument involves growing the cells on a thin, blackened polyester film which can be cut by the argon laser beam. Cells selected for cloning are then circumscribed. The heat of cutting welds the circumscribed film to a plastic coverslip surface or tissue culture chamber bottom. Nonselected cells may be removed by pulling the unattached polyester sheet from the attachment surface. The selected cells remain on polyester film disks welded to the plastic. Selections may be done automatically under computer control or manually by operator direction of stage movements. This instrument extends the art of automated cell selection and analysis to normal cell lines that must maintain cell‐substratum contact (anchorage dependence) for differentiated cell function, e.g., neurons, fibroblasts, or kidney cells.