The Cell Motility HCS Reagent Kit provides high-content screening (HCS)-quality fluorescence reagents and a protocol optimized for visualizing and quantifying cell movement by directly measuring the size of tracks generated by migrating cells. The kit provides a functional assay that can predict the efficacy of potential drugs for a number of therapeutic areas. The assay is performed on live cells plated on a lawn of microscopic fluorescent beads. As cells move across the lawn, they phagocytose the beads, clearing phagokinatic tracks behind them (Figure 1). The track area is proportional to the magnitude of cell movement. Included in the kit are buffers needed for processing the assay, blue fluorescent microspheres, and rhodamine-conjugated phaloidin, which stains f-actin, thus identifying the cell cytoplasm. Prepared cells can be analyzed using standard fluorescence microscopy or using a Cellomics fully automated HCS Readers and the Cell Motility BioApplication, affording automated plate handling, focusing, image acquisition, analysis, quantification and data storage.