Abstract
The resolution of fluorescence microscopy is limited by the diffraction imaging system, and many methods have been proposed to overcome the optical diffraction limit for achieving super-resolution imaging. Structured illumination microscopy (SIM) is one of the most competitive approaches and has demonstrated remarkable achievements. In the last two decades, SIM has been improved in many aspects, such as the enhancement of resolution and imaging depth and virtual modulation-based SIM. In this Perspective, we present an overview of the development of SIM, including the basic theory, application to biomedical studies, and the remarkable progress of SIM. Owing to its flexibility with respect to combination with other methods, SIM can be considered a powerful tool for biomedical study, offering augmented imaging capabilities by exploiting complementary advantages.
Highlights
Fluorescence microscopy is used extensively in biomedical research, providing several useful features, including minimal invasiveness, rapid data acquisition, targeting molecules of interest with specific labeling strategies, and real-time in vivo imaging.1 because of the optical diffraction limit, the highest attainable resolution of imaging is ∼200 nm.2 In diffractionlimited imaging systems, an infinitely small point is diffracted to a certain spot of finite size
In this Perspective, we present an overview of the development of Structured illumination microscopy (SIM), including the basic theory, application to biomedical studies, and the remarkable progress of SIM
Betzig et al proposed photoactivated localization microscopy (PALM),3 Rust et al described stochastic optical reconstruction microscopy (STORM),4 and Hess et al introduced fluorescence photoactivated localization microscopy (FPALM),5 of which the core principle of realizing super-resolution imaging is the control of fluorescent molecule emissions within the diffraction region at different times
Summary
Fluorescence microscopy is used extensively in biomedical research, providing several useful features, including minimal invasiveness, rapid data acquisition, targeting molecules of interest with specific labeling strategies, and real-time in vivo imaging. because of the optical diffraction limit (or Abbe diffraction limit), the highest attainable resolution of imaging is ∼200 nm. In diffractionlimited imaging systems, an infinitely small point is diffracted to a certain spot of finite size. Betzig et al proposed photoactivated localization microscopy (PALM), Rust et al described stochastic optical reconstruction microscopy (STORM), and Hess et al introduced fluorescence photoactivated localization microscopy (FPALM), of which the core principle of realizing super-resolution imaging is the control of fluorescent molecule emissions within the diffraction region at different times. Despite the remarkable resolution offered by SMLM, this approach is time consuming and requires specific fluorescent dyes for different biomedical imaging applications. Hell et al developed stimulated emission depletion microscopy (STED), which enables extremely fine control of the emission of fluorescence molecules by confining it to the central point within the diffraction region using high-intensity ring-shaped light, reducing the effective size of the diffraction spot significantly relative to conventional microscopy.. The challenges and solutions of SIM are discussed, in which SIM successfully augments its resolution, imaging depth, and virtual modulation-based SIM by combining SIM with different kinds of techniques
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