Research related to the development and application of luminescent nanoparticles (LNPs) for chemical and biological analysis and imaging is flourishing. Novel materials and new applications continue to be reported after two decades of research. This review provides a comprehensive and heuristic overview of this field. It is targeted to both newcomers and experts who are interested in a critical assessment of LNP materials, their properties, strengths and weaknesses, and prospective applications. Numerous LNP materials are cataloged by fundamental descriptions of their chemical identities and physical morphology, quantitative photoluminescence (PL) properties, PL mechanisms, and surface chemistry. These materials include various semiconductor quantum dots, carbon nanotubes, graphene derivatives, carbon dots, nanodiamonds, luminescent metal nanoclusters, lanthanidedoped upconversion nanoparticles and downshifting nanoparticles, triplet−triplet annihilation nanoparticles, persistent-luminescence nanoparticles, conjugated polymer nanoparticles and semiconducting polymer dots, multi-nanoparticle assemblies, and doped and labeled nanoparticles, including but not limited to those based on polymers and silica. As an exercise in the critical assessment of LNP properties, these materials are ranked by several application-related functional criteria. Additional sections highlight recent examples of advances in chemical and biological analysis, point-of-care diagnostics, and cellular, tissue, and in vivo imaging and theranostics. These examples are drawn from the recent literature and organized by both LNP material and the particular properties that are leveraged to an advantage. Finally, a perspective on what comes next for the field is offered.