Adaptive optics for fluorescence wide-field microscopy using spectrally independent guide star and markers

Vermeulen, Pierre; Muro, Eleonora; Pons, Thomas; Loriette, V.; Fragola, Alexandra

doi:10.1117/1.3603847

Cited by 20 publications

(11 citation statements)

References 22 publications

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“…Deformable mirrors are predominantly used in these applications as the detected light is typically broadband and randomly polarized. This is the case in transmission microscopes 37 and fluorescence widefield systems, 32,33,44,51 although an AO transmission microscope has been demonstrated using a SLM and narrowband light-emitting diode illumination. 52 Widefield imaging has also been performed in combination with adaptively corrected spatiotemporal focusing illumination.…”

Section: Widefield Microscopesmentioning

confidence: 99%

“…12,13 Another approach involves the use of point-like fluorescent objects as 'guide-stars' for the wavefront sensor. This can be implemented with fluorescent beads, 6,32,33 although this is not compatible with many bio-imaging applications. A more sophisticated approach relies upon the labelling of sparse structures, such as centrosomes, so that an array of suitable sensing sources is available, throughout a specimen.…”

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confidence: 99%

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Adaptive optical microscopy: the ongoing quest for a perfect image

2014

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Adaptive optics is becoming a valuable tool for high resolution microscopy, providing correction for aberrations introduced by the refractive index structure of specimens. This is proving particularly promising for applications that require images from deep within biological tissue specimens. We review recent developments in adaptive microscopy, including methods and applications. A range of advances in different microscope modalities is covered and prospects for the future are discussed. INTRODUCTIONOptical microscopes have for a long time played a vital role in biomedical research. Their ability to provide multidimensional structural and functional information about a specimen in a non-invasive manner has permitted numerous scientific advances. These microscopes rely upon high-quality optical systems to operate at the optimum resolution, which is determined by the diffraction limit of light for conventional microscopes. However, even with perfect optical components, the performance of the microscope is affected by the optical properties of the specimen. Spatial variations in refractive index introduce aberrations as the light passes through the specimen-a problem that is exacerbated when imaging deeps into tissue. These aberrations detrimentally affect the resolution and contrast of microscope images, ultimately limiting the depth at which imaging is practical. Adaptive optics (AO) was introduced into microscopy in order to overcome this limitation. Dynamic correction elements, such as deformable mirrors (DMs) or spatial light modulators (SLMs) have been employed to compensate specimen-induced aberrations (Figure 1). New wavefront sensing and control schemes have been developed to adapt the correction elements.The first demonstrations of adaptive optical microscopy took place in the early 2000s. In the few years that followed, there were several innovative advances that set the scene for further research. The research in this period is well covered by two review articles 1,2 and a recent compiled volume, 3 so this will not be reproduced here. This present article provides a review of advances in aberration correction in adaptive microscopy that have taken place in the last half-decade. During this period, a wide range of AO techniques have emerged, which have considerably enhanced the available toolkit. Furthermore, the benefit of correction of specimen-induced aberrations has been shown in different areas, particularly (although not exclusively) in biomedical applications. We outline these technological

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Section: Widefield Microscopesmentioning

confidence: 99%

mentioning

confidence: 99%

Adaptive optical microscopy: the ongoing quest for a perfect image

2014

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“…All of these results used some form of image optimisation based algorithm in which an image metric (sharpness, brightness, contrast being typical examples) was maximised by running through a range of mirror shapes. Whilst this method clearly worked in the temporally slowly varying microscopic image, a method using a measurement of the actual wavefront aberration, and hence correction via a closed loop system, was first shown by [8] using a back scattered source as the guide star and subsequently a fluorescent beacon [9] and [10]. A complete review of the work undertaken is well beyond the scope of this paper but the full range of methods and variations employed can be found in [11] where the background to image aberrations in optical microscopes is well explained along with an outline of the techniques employed and [12], which covers the practical implementation in particular in complex biological samples.…”

Section: Introductionmentioning

confidence: 99%

Comparison of closed loop and sensorless adaptive optics in widefield optical microscopy

Bourgenot

Saunter

Love

et al. 2013

J. Eur. Opt. Soc.-Rapid Publ.

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We report on a closed loop widefield adaptive optics, optical microscopy system in which the feedback signal is provided by backscattered light from the sample acting as a guide star. The improvement in imaging performance is compared to an adaptive optics system controlled via an image optimisation routine commonly described as sensorless adaptive optics. The samples viewed were imaged without fluorescence to ensure that photobleaching and other potential variations did not affect the comparisons in system performance though the method is equally applicable for fluorescence microscopy. The closed loop system is self-optimising for different areas of the sample, using a common reference wavefront, with the accuracy of the loop being limited by variation across the sub-aperture images induced by guide star elongation. Optimisation using an image sharpness metric gives slightly sharper images but takes significantly longer. We thus believe that both wavefront sensor based closed loop AO and metric based optimisation have a role to play in AO for microscopy and that the method of backscattered light as a guide star has a great potential in the application of AO, particularly to optical coherence tomography.

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“…This works extremely well, but is relatively slow and cannot easily be used on dynamic samples. More recent work has been published on full closed loop AO [48,61,42,50,51]. A major challenge for AO microscopy is obtaining a good signal for the WFS.…”

Section: Introductionmentioning

confidence: 99%

Light sheet adaptive optics microscope for 3D live imaging

Bourgenot

Taylor

Saunter

et al. 2013

Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XX

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The full-text may be used and/or reproduced, and given to third parties in any format or medium, without prior permission or charge, for personal research or study, educational, or not-for-prot purposes provided that:• a full bibliographic reference is made to the original source• a link is made to the metadata record in Durham E-Theses• the full-text is not changed in any wayThe full-text must not be sold in any format or medium without the formal permission of the copyright holders. DeclarationThe work in this thesis is based on research carried out at

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Adaptive optics for fluorescence wide-field microscopy using spectrally independent guide star and markers

Cited by 20 publications

References 22 publications

Adaptive optical microscopy: the ongoing quest for a perfect image

Adaptive optical microscopy: the ongoing quest for a perfect image

Comparison of closed loop and sensorless adaptive optics in widefield optical microscopy

Light sheet adaptive optics microscope for 3D live imaging

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