Parallel illumination for depletion microscopy through acousto-optic spatial light modulation

Abstract

Several types of super-resolution microscopy techniques, such as Stimulated Emission Depletion (STED), Reversible Saturable Optical Fluorescence Transitions (RESOLFT) or Switching Laser Mode (SLAM) microscopies, employ Laguerre-Gaussian beams (also called vortex or doughnut beams) to obtain fluorescence information within a sub-wavelength region of the specimen under observation, thus breaking the diffraction limit and producing images of greatly improved quality. However, in general, these techniques operate on a point-by-point basis, so scanning the sample in order to build a full, meaningful image, takes time. Parallelization of the illumination is the only way to make these microscopy techniques more suitable for real-time live cell imaging applications. Here, we demonstrate the parallel production of arbitrary arrays of Gaussian and Laguerre-Gaussian lasers foci suitable for super-resolution microscopy, together with the possibility to fast scan through the sample, by means of acousto-optic spatial light modulation, a technique that we have pioneered in the past in several other fields. Parallelized illumination with both Gaussian and doughnut beams will be then used to acquire super-resolution images.