Introduction to super-resolution microscopy

Yamanaka, Masahito; Smith, Nicholas I.; Fujita, Katsumasa

doi:10.1093/jmicro/dfu007

Cited by 102 publications

(47 citation statements)

References 105 publications

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“…The recent advent of superresolution microscopy introduced methods that surpass the diffraction limit of fluorescence imaging 16,17 and enabled experimental approaches that combine the high resolution previously exclusive to EM with the versatility of fluorescence microscopy 18 . SMLM is one of the main superresolution approaches, and it relies on determining the precise position of isolated images of single fluorophores 19 .…”

Section: Introductionmentioning

confidence: 99%

Correlated confocal and super-resolution imaging by VividSTORM

et al. 2015

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Single-molecule localization microscopy (SMLM) is rapidly gaining popularity in the life sciences as an efficient approach to visualize molecular distribution with nanoscale precision. However, it has been challenging to obtain and analyze such data within a cellular context in tissue preparations. Here we describe a 5-d tissue processing and immunostaining procedure that is optimized for SMLM, and we provide example applications to fixed mouse brain, heart and kidney tissues. We then describe how to perform correlated confocal and 3D-superresolution imaging on these sections, which allows the visualization of nanoscale protein localization within labeled subcellular compartments of identified target cells in a few minutes. Finally, we describe the use of VividSTORM (http://katonalab.hu/index.php/vividstorm), an open-source software for correlated confocal and SMLM image analysis, which facilitates the measurement of molecular abundance, clustering, internalization, surface density and intermolecular distances in a cell-specific and subcellular compartment-restricted manner. The protocol requires only basic skills in tissue staining and microscopy.

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Section: Introductionmentioning

confidence: 99%

Correlated confocal and super-resolution imaging by VividSTORM

et al. 2015

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“…In the current literature, there are articles reviewing optical sectioning, superresolution, or both [11][12][13][14][15][16][17][18][19][20][21][22][23]. Most of them treated structured illumination as one of the many techniques.…”

Section: Introductionmentioning

confidence: 99%

Structured illumination microscopy

2015

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Illumination plays an important role in optical microscopy. Köhler illumination, introduced more than a century ago, has been the backbone of optical microscopes. The last few decades have seen the evolution of new illumination techniques meant to improve certain imaging capabilities of the microscope. Most of them are, however, not amenable for wide-field observation and hence have restricted use in microscopy applications such as cell biology and microscale profile measurements. The method of structured illumination microscopy has been developed as a wide-field technique for achieving higher performance. Additionally, it is also compatible with existing microscopes. This method consists of modifying the illumination by superposing a well-defined pattern on either the sample itself or its image. Computational techniques are applied on the resultant images to remove the effect of the structure and to obtain the desired performance enhancement. This method has evolved over the last two decades and has emerged as a key illumination technique for optical sectioning, super-resolution imaging, surface profiling, and quantitative phase imaging of microscale objects in cell biology and engineering. In this review, we describe various structured illumination methods in optical microscopy and explain the principles and technologies involved therein.

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“…As a result, it is important for the spatial resolutions of two-photon excitation microscopes to be improved. Recently, several super-resolution imaging techniques have been developed [7]. Structured illumination microscopy (SIM) is a useful technique to maximize the frequency capabilities of imaging optics [8].…”

Section: Introductionmentioning

confidence: 99%

High-resolution imaging in two-photon excitation microscopy using in situ estimations of the point spread function

Doi¹,

Oketani²,

Nawa³

et al. 2017

Biomed. Opt. Express

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We present a technique for improving the spatial resolution of two-photon excitation microscopy; our technique combines annular illumination with an in situ estimation of the point spread function (PSF) used for deconvolution. For the in situ estimation of the PSF, we developed a technique called autocorrelation scanning, in which a sample is imaged by the scanning of two excitation foci that are overlapped over various distances. The image series obtained with the variation of the distance between the two foci provides the autocorrelation function of the PSF, which can be used to estimate the PSF at specific positions within a sample. We proved the principle and the effectiveness of this technique through observations of a fluorescent biological sample, and we confirmed that the improvement in the spatial resolution was ~1.7 times that of typical two-photon excitation microscopy by observing a mouse brain phantom at a depth of 200 µm.

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Introduction to super-resolution microscopy

Cited by 102 publications

References 105 publications

Correlated confocal and super-resolution imaging by VividSTORM

Correlated confocal and super-resolution imaging by VividSTORM

Structured illumination microscopy

High-resolution imaging in two-photon excitation microscopy using in situ estimations of the point spread function

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