Confocal microscopy has been widely used to acquire optical sectioning fluorescent image. However, traditional confocal technique requires point-by-point scanning which is time consuming. Alternative techniques to confocal microscopy, such as structured illumination, exist for fast sectioning images, but they require multiple axial planes to be imaged individually. Here, a non-axial line-scanning multifocal confocal microscopy is presented. The proposed system incorporates multiplex volume holographic grating (MVHG) in illumination and combination of multifocal image system. The detailed explanation for resolution on depth axial and simulation results are compared. Also both XY resolution is verified through resolution target. The ability of the proposed system to optical sectioning and multi-depth resolve image of fluorescently labeled microsphere and cornea is experimentally demonstrated.
Under broadband illumination, a multiplane microscopy incorporating volume holographic gratings (VHGs) to observe three-dimensional structures of biological samples with different depth simultaneously is presented. VHGs formed in thick recording materials, including PQ-PMMA, provide strong angular and wavelength transmittance filtering properties, which enable acquiring spatial–spectral images of fine structures within biological samples. Here, we experimentally demonstrate this microscopic imaging capability to obtain multiple depth-resolved mixed pollen grains images of fine structures from eight depths in one shot.