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High Content Screening (HCS) is a method for analyzing large numbers of cells typically arranged in arrays. Multi-well assays and micro-pallet arrays can be used to single out and harvest cells with desired features. However, visual inspection of large time series of images, each containing hundreds of wells or tens of thousands of micro-pallets is time-consuming and prone to detection errors.We have developed cell Analyst, a publicly available software package for automated identification and counting of cells. In this article we outline the image processing methods behind cell Analyst and show how this software can be applied to conduct highly specific cell counting.