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Fluorescence-based assays are widely used in high-throughput screening due to their high sensitivity, diverse selection of fluorophores, ease of operation, and various readout modes. As a result, fluorescence-based assays have been applied to monitor a broad range of activities in life-science research such as molecular dynamics and interactions, enzymatic activities, signal transduction, cell health, and distribution of molecules, organelles, or cells. This chapter describes two fluorescence-based techniques: total intensity measurement as an indication of cell viability, and fluorescence resonance energy transfer as an indication of protein folding and interactions, to illustrate in detail applications suitable for cell-based high-throughput screening in plate-reader and automated microscope-based formats, respectively.