Moreover, there have been numerous attempts to exploit SIM to achieve different goals. The axial resolution of SIM can be as good as that of the confocal microscope. Simple processing of wide-field images acquired with three lateral shifted patterns permits an optically sectioned image to be extracted in real-time. As a result, the microscope images efficiently only the portion of object where the fringe pattern is focused in. By projecting a spatial fringe pattern onto an object, only the zero spatial frequency does not attenuate with defocus. 16 as a means to eliminate the out-of-focus background encountered in wide-field microscopy and to improve its signal-to-noise ratio. A number of far-field optical super-resolution microscopic techniques, such as Stimulated Emission Depletion (STED) microscopy 1, 2, 3, Structured Illumination Microscopy (SIM) 4, 5, Stochastic Optical Reconstruction Microscopy (STORM) 6, 7, Photo-Activated Localization Microscopy (PALM) 8 and so forth have since been developed.Īs a wide-field optical microscopy, SIM has found widespread applications for investigations of subcellular structures 9, 10, 11 and for time-series imaging of living cells 12, 13, 14, 15 due to its resolution beyond the diffraction-limit, its axial sectioning capability, as well as its high imaging speed. Over the past decade, this limitation has driven the exploration of super-resolution optical imaging methodologies. This is larger than many subcellular structures to be resolved. However, the spatial resolution of conventional far-field optical microscopy is limited to about 200 nm, due to the well-known Abbe diffraction-limit. The key advantages, especially of far-field optical microscopy, over other forms of microscopy are the capability and compatibility of non-contact, minimally invasive observation and measurement of live specimens. The 2D super-resolution and 3D optical sectioning modalities can be easily switched and applied to either fluorescent or non-fluorescent specimens.ĭespite of the availability of various high resolution microscopic technologies, e.g., scanning electron microscopy (SEM), scanning tunnel microscopy (STM), atomic force microscopy (AFM), optical microscopy plays an essential role in biological studies. 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 maximum acquisition speed for 3D imaging in the optical sectioning mode was 1.6×10 7 pixels/second, which was mainly limited by the sensitivity and speed of the CCD camera. The lateral resolution of 90 nm and the optical sectioning depth of 120 μm were achieved. 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. 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.