Single-particle tracking of immunoglobulin E receptors (FcεRI) in micron-sized clusters and receptor patches

When mast cells contact a monovalent antigen-bearing fluid lipid bilayer, IgE-loaded FcεRI receptors aggregate at contact points and trigger degranulation and the release of immune activators. We used two-color total internal reflection fluorescence microscopy and single-particle tracking to show that most fluorescently labeled receptor complexes diffuse freely within these micron-size clusters, with a diffusion coefficient comparable to free receptors in resting cells. At later times, when the small clusters coalesce to form larger patches, receptors diffuse even more rapidly. In all cases, Monte Carlo diffusion simulations ensured that the tracking results were free of bias, and distinguished biological from statistical variation. These results show the diversity in receptor mobility in mast cells, demonstrating at least three distinct states of receptor diffusivity.

► Super-resolution single-particle tracking of immune receptors.
► FcεRI receptors maintain lateral mobility inside receptor clusters.
► FcεRI receptor immobilization is not required for signaling.
► Monte Carlo calculations distinguish biological from statistical variability.