In-depth single molecule localization microscopy using adaptive optics and single objective light-sheet microscopy

Single molecule localization microscopy (SMLM) allows deciphering the nanoscale organization and dynamics of biomolecules in their native environment with unprecedented resolution. While SMLM was quickly adopted by the scientific community for its performance and simple instrumentation, it still remains limited in its in-depth capability, precluding many biological processes to be investigated. We here present a solution to perform in-depth volumetric SMLM, called soSMARt. It relies on innovative microfabricated devices allowing both single-objective light-sheet microscopy, aberrations correction via adaptive optics, and real-time feedback-loop registration with nanometric precision. We illustrated the performances of soSMARt to assess the 3D nanoscale organization of several protein of interest in isolated cells, and explore optimizations and proof-of-concepts for the investigation of more complex tissues such as 3D cell cultures. We believe our method addresses key limitations of single molecule microscopy, paving the way for novel biological applications.