Extended depth-of-field microscopy with a high-speed deformable mirror

Shain, William J.; Vickers, Nicholas A.; Goldberg, Bennett B.; Bifano, Thomas G.; Mertz, Jérôme

doi:10.1364/ol.42.000995

Cited by 120 publications

(76 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…As expected, when the acoustic lens was turned on ( Figure 3A), the excitation beam was scanned multiple times over the z-axis at each pixel, and consequently, the integration of the pixel information resulted in an extension in the axial PSF from 1.5 μm to about 30 μm (FWHM values). Note that the achieved axial scanning range was in agreement with the theoretical value of 35 μm [30,45]. Importantly, the lateral PSF throughout the whole scanning range (average FWHM = 0.60 μm) was comparable to the in-focus PSF with lens off (FWHM ' 0.55 μm) ( Figure 3B,C).…”

Section: Fast Volumetric Imagingsupporting

confidence: 85%

Enhanced volumetric imaging in 2‐photon microscopy via acoustic lens beam shaping

Piazza

Bianchini

Sheppard

et al. 2017

Journal of Biophotonics

View full text Add to dashboard Cite

Three-dimensional imaging at high-spatiotemporal resolutions and over large penetration depths is key for unmasking the dynamics and structural organization of complex biological systems. However, the need to axially shift the focus, with consequent limitations in imaging speed, and signal degradation at large depths due to scattering effects, makes this task challenging. Here, we present a novel approach in 2-photon excitation microscopy that allows fast volumetric imaging and enhanced signal-to-background (S/B) in thick tissue. Our technique is based on ultrafast beam shaping at each pixel by means of an acoustic optofluidic lens. Shaping the excitation beam with different phase profiles enables both high-speed axial focus shifting, for continuous volumetric imaging, and controlled aberrated imaging, advantageous for out-of-focus background removal. We provide a theoretical description of our approach, and demonstrate volumetric imaging of neuronal cells from a mouse brain slice with enhancements in S/B up to a factor of 10 over a depth of 600 μm.

show abstract

Section: Fast Volumetric Imagingsupporting

confidence: 85%

Enhanced volumetric imaging in 2‐photon microscopy via acoustic lens beam shaping

Piazza

Bianchini

Sheppard

et al. 2017

Journal of Biophotonics

View full text Add to dashboard Cite

show abstract

“…While lateral 2D-scans can achieve video rate by employing resonant scanners or acousto-optic deflectors, a sufficiently fast modulation of the axial focus position is still challenging. Partly in response to this specific demand, in recent years several technologies have been developed to tune the optical power of lenses (Berge, 2005;Ren et al, 2006;Bernet et al, 2013) or Micro-Electro-Mechanical System mirrors (Qi et al, 2004;Shain et al, 2017). Some of these systems, like the tunable polymer-filled lenses (Ren et al, 2006;Jabbour et al, 2014;Nakai et al, 2015) or lenses based on electrowetting (Berge, 2005) are now available.…”

Section: Introductionmentioning

confidence: 99%

Remote focusing in confocal microscopy by means of a modified Alvarez lens

Bawart

Jesacher

Bernet

et al. 2018

Journal of Microscopy

View full text Add to dashboard Cite

Alvarez lenses are actuated lens-pairs which allow one to tune the optical power by mechanical displacement of subelements. Here, we show that a recently realized modified Alvarez lens design which does not require mechanical actuation can be integrated into a confocal microscope. Instead of mechanically moving them, the sublenses are imaged onto each other in a 4f-configuration, where the lateral image shift leading to a change in optical power is created by a galvo-mirror. The avoidance of mechanical lens shifts leads to a large speed gain for axial (and hence also 3D) image scans compared to classical Alvarez lenses. We demonstrate that the suggested operation principle is compatible with confocal microscopy. In order to optimize the system, we have drawn advantage of the flexibility a liquid-crystal spatial light modulator offers for the implementation. For given specifications, dedicated diffractive optical elements or freeform elements can be used in combination with resonant galvo-scanners or acousto-optic beam deflectors, to achieve even faster z-scans than reported here, reaching video rate.

show abstract

“…The core of our technique involves sweeping the focal plane of a microscope at an axial position z s over a scan range D. There are many ways of implementing a focal sweep system [15]; while any such system would be acceptable, our method is to insert a deformable mirror (DM) into the back focal plane, or pupil plane, of an otherwise standard epi-fluorescence microscope (described in [22]). The curvature of the DM can be swept from positive to negative, causing the focal plane of the microscope to sweep over a scan range from z s = − D 2 to z s = D 2 , during a single camera exposure.…”

Section: Introductionmentioning

confidence: 99%

“…The curvature of the DM can be swept from positive to negative, causing the focal plane of the microscope to sweep over a scan range from z s = − D 2 to z s = D 2 , during a single camera exposure. The key benefits of using a DM are light efficiency, achromaticity, speed (we use a DM that provides a 20kHz update rate), and scan range (we have shown that even with a modest DM stroke of 2.5 − 3µm, we can extend the standard depth of field of the microscope by ≈ 70× [22]). In existing focal-sweeping systems, the EDOF image is a single-shot projection of Fig.…”

Section: Introductionmentioning

confidence: 99%

Axial localization with modulated-illumination extended-depth-of-field microscopy

Shain

Vickers

et al. 2018

Biomed. Opt. Express

Self Cite

View full text Add to dashboard Cite

High-speed volumetric imaging represents a challenge in microscopy applications. We demonstrate a technique for acquiring volumetric images based on the extended depth of field microscopy with a fast focal scan and modulated illumination. By combining two frames with different illumination ramps, we can perform local depth ranging of the sample at speeds of up to half the camera frame rate. Our technique is light efficient, provides diffraction-limited resolution, enables axial localization that is largely independent of sample size, and can be operated with any standard widefield microscope based on fluorescence or darkfield contrast as a simple add-on. We demonstrate the accuracy of axial localization and applications of the technique to various dynamic extended samples, including in-vivo mouse brain.

show abstract

Extended depth-of-field microscopy with a high-speed deformable mirror

Cited by 120 publications

References 25 publications

Enhanced volumetric imaging in 2‐photon microscopy via acoustic lens beam shaping

Enhanced volumetric imaging in 2‐photon microscopy via acoustic lens beam shaping

Remote focusing in confocal microscopy by means of a modified Alvarez lens

Axial localization with modulated-illumination extended-depth-of-field microscopy

Contact Info

Product

Resources

About