This volume explores the latest available wet-lab techniques and computational methods to study in-cell small-molecule behavior and interactions with their targets. The chapters in this book discuss topics such as disease-relevant models for chemical biology studies, target engagement using cellular thermal shift assay or bioluminescence resonance energy transfer; visualization of bio-active small molecules Raman microscopy; (phospho-)proteomics and transcriptomics for mode-of-action studies, CRISPR/Cas9-based chemogenomic profiling in mammalian cells; predicting drug interactions using computational approaches; comparison of compound-induced profiles using high-content imaging or cancer cell line panels and web-based tools for polypharmacology prediction. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Cutting-edge and thorough, Systems Chemical Biology: Methods and Protocols is a valuable resource for novice or expert scientists and researchers trying to initiate or continue their chemical biology studies at a systems level.