Nonlinear super-resolution imaging with adaptive optical multifocal structured illumination microscopy

Optical microscopy has been indispensable for physiologically visualizing neuronal structure and function, while it remains a challenge since the limited diffraction resolution and restricted imaging depth. Nonlinear multifocal structured illumination microscopy (MSIM) provides resolution-doubled images and good penetration, but its performance is seriously degraded in deep tissues. Thus, we present a non-inertial scanning nonlinear MSIM system combined with adaptive optics (AO) to realize super-resolution imaging with aberration correction in vivo. It is indispensable for visualizing various synaptic spines and detailed morphological changes of neurons in vivo. Using the nonlinear MSIM for SHG imaging, collagen fiber alignments were quantified in various tissues, demonstrating its feasibility for identifying collagen characteristics.