Structured illumination microscopy (SIM), as one of the mainstream super-resolution optical microscopy imaging techniques, takes the advantages of fast imaging because of wide-field illumination, which is suitable to the multi-color live cell imaging. However, multi-color imaging measures each spectral image separately, so the imaging speed of is limited. In addition, the spatial-resolution improvement of SIM depends on the modulation frequency of structured pattern. To further improve the imaging resolution, we use a spectral imaging method, i.e., passive ghost imaging, in SIM. Simulation results show that the proposed method has superiority in super-resolution, imaging speed and imaging quality than traditional multi-color imaging and multi-color SIM imaging.
SignificanceDouble-helix point spread function (DH-PSF) microscopy has been developed for three-dimensional (3D) localization and imaging at super-resolution but usually in environments with no or weak scattering. To date, super-resolution imaging through turbid media has not been reported.AimWe aim to explore the potential of DH-PSF microscopy in the imaging and localization of targets in scattering environments for improved 3D localization accuracy and imaging quality.ApproachThe conventional DH-PSF method was modified to accommodate the scanning strategy combined with a deconvolution algorithm. The localization of a fluorescent microsphere is determined by the center of the corresponding double spot, and the image is reconstructed from the scanned data by deconvoluting the DH-PSF.ResultsThe resolution, i.e., the localization accuracy, was calibrated to 13 nm in the transverse plane and 51 nm in the axial direction. Penetration thickness could reach an optical thickness (OT) of 5. Proof-of-concept imaging and the 3D localization of fluorescent microspheres through an eggshell membrane and an inner epidermal membrane of an onion are presented to demonstrate the super-resolution and optical sectioning capabilities.ConclusionsModified DH-PSF microscopy can image and localize targets buried in scattering media using super-resolution. Combining fluorescent dyes, nanoparticles, and quantum dots, among other fluorescent probes, the proposed method may provide a simple solution for visualizing deeper and clearer in/through scattering media, making in situ super-resolution microscopy possible for various demanding applications.
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