Super-resolved live-cell imaging using random illumination microscopy
Thomas Mangeat, Simon Labouesse, Marc Allain, Awoke Negash, Emmanuel Martin, Aude Guénolé, Renaud Poincloux, Claire Estibal, Anaïs Bouissou, Sylvain Cantaloube, Elodie Vega, Tong Li, Christian Rouvière, Sophie Allart, Debora Keller, Valentin Debarnot, Xia Bo Wang, Grégoire Michaux, Mathieu Pinot, Roland Le Borgne, Sylvie Tournier, Magali Suzanne, Jérôme Idier, Anne Sentenac
Abstract
Current super-resolution microscopy (SRM) methods suffer from an intrinsic complexity that might curtail their routine use in cell biology. We describe here random illumination microscopy (RIM) for live-cell imaging at super-resolutions matching that of 3D structured illumination microscopy, in a robust fashion. Based on speckled illumination and statistical image reconstruction, easy to implement and user-friendly, RIM is unaffected by optical aberrations on the excitation side, linear to brightness, and compatible with multicolor live-cell imaging over extended periods of time. We illustrate the potential of RIM on diverse biological applications, from the mobility of proliferating cell nuclear antigen (PCNA) in U2OS cells and kinetochore dynamics in mitotic S. pombe cells to the 3D motion of myosin minifilaments deep inside Drosophila tissues. RIM's inherent simplicity and extended biological applicability, particularly for imaging at increased depths, could help make SRM accessible to biology laboratories.