DMD-based LED-illumination Super-resolution and optical sectioning microscopy

Abstract

Super-resolution three-dimensional (3D) optical microscopy has incomparable advantages over other high-resolution microscopic technologies, such as electron microscopy and atomic force microscopy, in the study of biological molecules, pathways and events in live cells and tissues. We present a novel approach of structured illumination microscopy (SIM) by using a digital micromirror device (DMD) for fringe projection and a low-coherence LED light for illumination. The lateral resolution of 90 nm and the optical sectioning depth of 120 μm were achieved. The maximum acquisition speed for 3D imaging in the optical sectioning mode was 1.6×107 pixels/second, which was mainly limited by the sensitivity and speed of the CCD camera. In contrast to other SIM techniques, the DMD-based LED-illumination SIM is cost-effective, ease of multi-wavelength switchable and speckle-noise-free. The 2D super-resolution and 3D optical sectioning modalities can be easily switched and applied to either fluorescent or non-fluorescent specimens.

Single-pixel imaging by means of Fourier spectrum acquisition

“…The proposed technique employs the use of greyscale, analytic, harmonic sinusoid patterns (the real part of Fourier transform basis), instead of random speckle or binary basis patterns, for illumination. There are approaches to super-resolution microscopy with the use of the sinusoid structured illumination [14][15][16] . The proposed technique acquires the Fourier spectrum of the scene image with the fourstep phase-shifting sinusoid illuminations.…”

“…In addition, the sinusoid patterns can be generated by several means. For example, a commercial digital projector displays the patterns in 24-bit mode; a programmable digital micromirror device (DMD) that displays the patterns in binary mode, and the defocusing 17 or the spatial filtering approach 16 is employed to make the resulting patterns sinusoidal; two coherent plane waves intersect at an angle to form a sinusoid pattern. What is more, the technique presented here is a compressive sampling 4,[8][9][10]12,13 like approach.…”

Super-resolution microscopy and its applications in neuroscience

“…The excitation beam exhibits a periodically spatial pattern, 8 which is originally a sinusoidal grid that can be generated by a digital micromirror device, 40 replacing the conventional uniform illumination. The spatial frequency of the excitation light mixes with the specimen, shifting high frequency information of the sample into lower frequency which then could be detected by the microscope.…”

Super-resolution fluorescence polarization microscopy

“…Super-resolution techniques break the di®raction barrier, which is achieved through intensity on-o® modulation. The modulation could be in a structured manner, such as stimulated emission depletion (STED) 54,55 or (saturated) structured illumination microscopy (SIM), [56][57][58] or in a stochastic manner, such as (f) photo-activated localization microscopy (PALM)/ stochastic optical reconstruction microscopy (STORM). [59][60][61][62] Though a large variety of super resolution techniques have been developed for°uo-rescence intensity imaging, super resolution FPM is just starting.…”