A guided tour into subcellular colocalization analysis in light microscopy


  • S. BOLTE,

    1. Plateforme d’Imagerie et de Biologie Cellulaire, IFR 87 ‘la Plante et son Environnement’, Institut des Sciences du Végétal, Avenue de la Terrasse, 91198 Gif-sur-Yvette Cedex, France
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    1. Institut Curie, CNRS UMR 146, Plateforme d’Imagerie Cellulaire et Tissulaire, Bâtiment 112, Centre Universitaire, 91405 Orsay Cedex, France
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S. Bolte. Tel: 0033 69863130; Fax: 0033 169 86 1703; e-mail: Susanne.Bolte@isv.cnrs-gif.fr.
F. P. Cordelières. E-mail: Fabrice.Cordelieres@curie.u-psud.fr


It is generally accepted that the functional compartmentalization of eukaryotic cells is reflected by the differential occurrence of proteins in their compartments. The location and physiological function of a protein are closely related; local information of a protein is thus crucial to understanding its role in biological processes. The visualization of proteins residing on intracellular structures by fluorescence microscopy has become a routine approach in cell biology and is increasingly used to assess their colocalization with well-characterized markers. However, image-analysis methods for colocalization studies are a field of contention and enigma. We have therefore undertaken to review the most currently used colocalization analysis methods, introducing the basic optical concepts important for image acquisition and subsequent analysis. We provide a summary of practical tips for image acquisition and treatment that should precede proper colocalization analysis. Furthermore, we discuss the application and feasibility of colocalization tools for various biological colocalization situations and discuss their respective strengths and weaknesses. We have created a novel toolbox for subcellular colocalization analysis under ImageJ, named JACoP, that integrates current global statistic methods and a novel object-based approach.