This detailed collection explores recent advances in molecular imaging techniques involving bioluminescence, currently employed in biolaboratories around the world. Volume 1 delves into techniques for the establishment of luciferins and luciferases, basic in vitro and in vivo applications, as well as protocols on multiplex imaging platforms. Written for the highly successful Methods in Molecular Biology series, 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. Authoritative and comprehensive, Bioluminescence: Methods and Protocols, Fourth Edition, Volume 1 presents practical guidance for researchers and technical staff on how to proceed with bioluminescence studies in their laboratories.