title: Clus-DoC: a combined cluster detection and colocalization analysis for single-molecule localization microscopy data.
abstract: Advances in fluorescence microscopy are providing increasing evidence that the spatial organization of proteins in cell membranes may facilitate signal initiation and integration for appropriate cellular responses. Our understanding of how changes in spatial organization are linked to function has been hampered by the inability to directly measure signaling activity or protein association at the level of individual proteins in intact cells. Here we solve this measurement challenge by developing Clus-DoC, an analysis strategy that quantifies both the spatial distribution of a protein and its colocalization status. We apply this approach to the triggering of the T-cell receptor during T-cell activation, as well as to the functionality of focal adhesions in fibroblasts, thereby demonstrating an experimental and analytical workflow that can be used to quantify signaling activity and protein colocalization at the level of individual proteins.
authors: EMBL Australia Node in Single Molecule Science, School of Medical Sciences, and ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia.
authors: Sophie V
authors: SV
authors: Pageon
authors: EMBL Australia Node in Single Molecule Science, School of Medical Sciences, and ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia.
authors: Philip R
authors: PR
authors: Nicovich
authors: EMBL Australia Node in Single Molecule Science, School of Medical Sciences, and ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia.
authors: Mahdie
authors: M
authors: Mollazade
authors: EMBL Australia Node in Single Molecule Science, School of Medical Sciences, and ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia.
authors: Thibault
authors: T
authors: Tabarin
authors: EMBL Australia Node in Single Molecule Science, School of Medical Sciences, and ARC Centre of Excellence in Advanced Molecular Imaging, University of New South Wales, Sydney, NSW 2052, Australia k.gaus@unsw.edu.au.
authors: Katharina
authors: K
authors: Gaus
mesh_terms: D005347:Fibroblasts; D022001:Focal Adhesions; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D008213:Lymphocyte Activation; D008565:Membrane Proteins; D008856:Microscopy, Fluorescence / Q000379:methods; D011948:Receptors, Antigen, T-Cell; D015398:Signal Transduction; D000072760:Single Molecule Imaging / Q000379:methods*; D012984:Software; D062206:Spatial Analysis
chemical_list: D008565:Membrane Proteins; D011948:Receptors, Antigen, T-Cell