Cell Supply High Throughput and High Content Assays 127

Christa Burger, Oliver Poschke, and Mirek R. Jurzak

5.1 Introduction 127

5.2 Cell Lines for HTS 128

5.2.1 Selection of the Most Suitable Cell Line 128

5.2.2 Optimizing Cell Cultivation 130

5.2.2.1 Adherence 130

5.2.2.2 pH and Temperature 130

5.2.2.3 Media and Additives 131

5.2.2.4 Solvent Tolerance 131

5.2.2.5 Cell Density 131

5.2.3 Optimizing the Reproducibility of Seeding 132

5.2.3.1 Signal Shift 132

5.2.3.2 Edge Effect 132

5.2.4 Cell Production and Plate Delivery 132

5.2.4.1 The Amount of Cells Needed 132

5.2.4.2 Cell Storage 133

5.3 Conventional Cellular Screening Assays 134

5.3.1 General HTS Assay Prerequisites 134

5.3.2 Evaluation of Assay Quality 134

5.3.3 ELISA-Based Assays 135

5.3.4 Radiometric Cellular Assays 136

5.3.5 Reporter Gene Assays 137

5.3.6 Second Messenger Assays 138

5.3.7 Ion Channel Assays 138

5.4 The Definition of High-Content Screening 139

5.4.1 Instrumentation for HCS 139

5.4.2 Reagents (Fluorescent Probes) for HCS 140

5.4.2.1 Low-Molecular-Weight Fluorophores 140

5.4.2.2 Genetically Encoded Reporter for Fluorescence Detection 141

5.4.3 Assays and Target-Based Applications of HCS 142

5.4.3.1 GPCRs 142

5.4.3.2 Kinases 143

5.4.3.3 Other Drug Targets 144

5.4.4 HCS Applications Targeting Generic Cellular Parameters and Morphology 145

5.5 Outlook 146 References 147

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