MIiSR: Molecular Interactions in Super-Resolution Imaging Enables the Analysis of Protein Interactions, Dynamics and Formation of Multi-protein Structures
Fig 4
Quantifying co-clustering using the radial distribution function and Ripley’s K function.
(A) The β2-adrenergic receptor (β2-AR, Green), β-arrestin-1 (βarr1, Red) and Clathrin (Blue) form an internalization complex after stimulation of the β2-adrenergic receptor in HEK293 cells with 10 μM isoproterenol. (B-C) Quantification of co-clustering of the β2-adrenergic receptor with β-arrestin-1 by (D) RDF analysis and (E) Ripley’s H-function, before (0 min) and 1 and 5 minutes post-stimulation with isoproterenol. (D-E) Quantification of β2-adrenergic receptor co-clustering with clathrin coated pits by (D) RDF analysis and (E) Ripley’s H-function, before (0 min) and 1 and 5 minutes post-stimulation with isoproterenol. (F) Self-clustering of the tetraspanin CD81 induced by the use of polyvalent versus mono/divalent labeling reagents. Ab = antibody. All images are representative of, and graphs quantify, 3 experiments with a minimum of 5 images per experiment. Data are presented as mean ± SEM, * = p < 0.05, n.s. = not significantly different, ANOVA with Bonferroni correction. Scale bar (A) is 2.5 μm, inserts are 1.47 μm x 1.47 μm. D-E: Arrows indicate mean cluster size, as determined by the radius of maximum H(r).