biostudies-literatureEjdrup ALLundbeckfondenLundbeck Foundation4388https://www.ebi.ac.uk/biostudies/studies/S-EPMC9334352biostudies-literatureUnknown13(1)Dual-color single-molecule localization microscopy (SMLM) provides unprecedented possibilities for detailed studies of colocalization of different molecular species in a cell. However, the informational richness of the data is not fully exploited by current analysis tools that often reduce colocalization to a single value. Here, we describe a tool specifically designed for determination of co-localization in both 2D and 3D from SMLM data. The approach uses a function that describes the relative enrichment of one molecular species on the density distribution of a reference species. The function reframes the question of colocalization by providing a density-context relevant to multiple biological questions. Moreover, the function visualize enrichment (i.e. colocalization) directly in the imaNature communicationsA density-based enrichment measure for assessing colocalization in single-molecule localization microscopy data.PMC9334352R230-2016-3154R303-2018-3540R276-2018-792R266-2017-4331Madsen KLGether UKonomi AHerborg FEjdrup ALLycas MDLorenzen NfalseA density-based enrichment measure for assessing colocalization in single-molecule localization microscopy data.Dual-color single-molecule localization microscopy (SMLM) provides unprecedented possibilities for detailed studies of colocalization of different molecular species in a cell. However, the informational richness of the data is not fully exploited by current analysis tools that often reduce colocalization to a single value. Here, we describe a tool specifically designed for determination of co-localization in both 2D and 3D from SMLM data. The approach uses a function that describes the relative enrichment of one molecular species on the density distribution of a reference species. The function reframes the question of colocalization by providing a density-context relevant to multiple biological questions. Moreover, the function visualize enrichment (i.e. colocalization) directly in the ima2022-01-01T00:00:00Z2022 Jul2025-04-19T11:15:45.338Z2025-04-19T11:15:45.338ZS-EPMC93343523590257810.1038/s41467-022-32064-y