Revealing mitochondrial architecture and functions with single molecule localization microscopy

Understanding the spatiotemporal organization of components within living systems requires the highest resolution possible. Microscopy approaches that allow for a resolution below 250 nm include electron and super-resolution microscopy (SRM). The latter combines advanced imaging techniques and the optimization of image processing methods. Over the last two decades, various SRM-related approaches have been introduced, especially those relying on single molecule localization microscopy (SMLM). To develop and apply SMLM approaches, mitochondria are an ideal cellular compartment due to their size, which is below the standard diffraction limit. Furthermore, mitochondria are a dynamic yet narrow compartment, and a resolution below 250 nm is required to study their composition and multifaceted functions. To this end, several SMLM technologies have been used to reveal mitochondrial composition. However, there is still room for improvement in existing techniques to study protein-protein interactions and protein dynamics within this compartment. This review aims to offer an updated overview of the existing SMLM techniques and probes associated with mitochondria to enhance their resolution at the nanoscale. Last, it paves the way for future SMLM improvements to better resolve mitochondrial dynamics and functions.