Programmable Aperture Light‐Field Microscopy

Abstract A computational three‐dimensional (3D) microscopy technique, termed programmable aperture light‐field microscopy (PALFM), for motion‐free, high‐resolution volumetric imaging of fluorescent or self‐luminous samples is proposed. The well‐known Fourier slice theorem is extended to incoherent tomographic imaging, which states that a detected image under an ideal aperture corresponds to a central slice in the 3D object spectrum, so the spectrum coverage can be accomplished based on the motion‐free aperture modulation. When further considering frequency extension and coverage, a hybrid aperture modulation scheme is designed consisting of non‐centrosymmetric circular and annular apertures for high‐efficiency, non‐ambiguous depth discrimination. A PALFM system with an easy‐to‐build programmable aperture module attached to an off‐the‐shelf inverted fluorescence microscope is constructed, where annular apertures can manipulate Bessel‐like beams for spectrum modulation. Experimental results on near‐diffraction‐limited imaging of a resolution target across a large depth range and high‐resolution, multi‐color 3D imaging of a mouse kidney section verify the validity and effectiveness of PALFM. High‐speed, long‐term time‐lapse volumetric imaging of HeLa cells in vitro further demonstrates that PALFM is a promising tomographic imaging tool for studying dynamic cellular processes and events without requiring complicated sample rotation or beam scanning.