Widefield compressive multiphoton microscopy

Alemohammad, Milad; Shin, Jaewook; Trần, Dũng; Stroud, Jasper R.; Chin, Sang; Tran, Trac D.; Foster, Mark A.

doi:10.1364/ol.43.002989

Cited by 16 publications

(12 citation statements)

References 21 publications

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“…A complete imaging experiment with 1024 patterns can now be performed in 17 s. In addition, with the use of a state-of-the art DMD and compressive sensing, imaging speed could be vastly increased. 31 Sub-1 min imaging and recovery is feasible for image sampling and FOVs matching conventional microscopy.…”

Section: Discussionmentioning

confidence: 99%

Wide-field multiphoton imaging with TRAFIX

Escobet-Montalbán

Wijesinghe

Chen

et al. 2019

Multiphoton Microscopy in the Biomedical Sciences XIX

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Optical approaches have broadened their impact in recent years with innovations in both wide-field and superresolution imaging, which now underpin biological and medical sciences. Whilst these advances have been remarkable, to date, the ongoing challenge in optical imaging is to penetrate deeper. TRAFIX is an innovative approach that combines temporal focusing illumination with single-pixel detection to obtain wide-field multiphoton images of fluorescent microscopic samples deep through scattering media without correction. It has been shown that it can image through biological samples such as rat brain or human colon tissue up to a depth of seven scattering mean-free-path lengths. Comparisons of TRAFIX with standard point-scanning two-photon microscopy show that the former can yield a five-fold higher signal-to-background ratio while significantly reducing photobleaching of the specimen. Here, we show the first preliminary demonstration of TRAFIX with three-photon excitation imaging dielectric beads. We discuss the advantages of the TRAFIX approach combined with compressive sensing for biomedicine.

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Section: Discussionmentioning

confidence: 99%

Wide-field multiphoton imaging with TRAFIX

Escobet-Montalbán

Wijesinghe

Chen

et al. 2019

Multiphoton Microscopy in the Biomedical Sciences XIX

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“…Here, we demonstrate a powerful approach to "de-scatter" widefield TFM images. Many computational imaging approaches have previously been proposed (12)(13)(14)(15); while they improve image resolution and contrast at shallower depths (<1 scattering length), most of them have not been applied for deep imaging. Notably, Escobet-Montalbán and co-workers (9) demonstrated a method called "TempoRAl Focusing microscopy with single-pIXel detection (TRAFIX)"; they used a set of two-dimensional illumination patterns along with single pixel detection (16) to image as deep as 7 scattering lengths through a scattering phantom (9).…”

Section: Introductionmentioning

confidence: 99%

De-scattering with Excitation Patterning enables rapid wide-field imaging through scattering media

et al. 2021

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Nonlinear optical microscopy has enabled in vivo deep tissue imaging on the millimeter scale. A key unmet challenge is its limited throughput especially compared to rapid wide-field modalities that are used ubiquitously in thin specimens. Wide-field imaging methods in tissue specimens have found successes in optically cleared tissues and at shallower depths, but the scattering of emission photons in thick turbid samples severely degrades image quality at the camera. To address this challenge, we introduce a novel technique called De-scattering with Excitation Patterning or “DEEP,” which uses patterned nonlinear excitation followed by computational imaging–assisted wide-field detection. Multiphoton temporal focusing allows high-resolution excitation patterns to be projected deep inside specimen at multiple scattering lengths due to the use of long wavelength light. Computational reconstruction allows high-resolution structural features to be reconstructed from tens to hundreds of DEEP images instead of millions of point-scanning measurements.

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“…Widefield imaging has been successfully demonstrated for two-dimensional (2-D) twophoton fluorescence (TPF) and SHG microscopy, albeit at the expense of requiring a higher average illumination power than that used in point-scanning techniques and no intrinsic axial sectioning capability. [19][20][21][22] On this latter point, widefield illumination has either been combined with digital holography for three-dimensional (3-D) image reconstruction 23,24 or temporal focusing [25][26][27] to restrict the multiphoton effect to selected planes along the optical axis. In this paper, our analysis is performed in 2-D, which is sufficient for many practical applications that does not require 3-D reconstruction.…”

Section: Introductionmentioning

confidence: 99%

Demonstration of flat-top beam illumination in widefield multiphoton microscopy

et al. 2019

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Multiphoton microscopy provides a suitable technique for imaging biological tissues with submicrometer resolution. Usually a Gaussian beam (GB) is used for illumination, leading to a reduced power efficiency in the multiphoton response and vignetting for a square-shaped imaging area. A flat-top beam (FTB) provides a uniform spatial intensity distribution that equalizes the probability of a multiphoton effect across the imaging area. We employ a customized widefield multiphoton microscope to compare the performance of a squareshaped FTB illumination with that based on using a GB, for both two-photon fluorescence (TPF) and second-harmonic generation (SHG) imaging. The variation in signal-to-noise ratio across TPF images of fluorescent dyes spans ∼5.6 dB for the GB and ∼1.2 dB for the FTB illumination, respectively. For the GB modality, TPF images of mouse colon and Convallaria root, and SHG images of chicken tendon and human breast biopsy tissue showcase ∼20% area that are not imaged due to either insufficient or lack of illumination. For quantitative analysis that depends on the illuminated area, this effect can potentially lead to inaccuracies. This work emphasizes the applicability of FTB illumination to multiphoton applications.

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Widefield compressive multiphoton microscopy

Cited by 16 publications

References 21 publications

Wide-field multiphoton imaging with TRAFIX

Wide-field multiphoton imaging with TRAFIX

De-scattering with Excitation Patterning enables rapid wide-field imaging through scattering media

Demonstration of flat-top beam illumination in widefield multiphoton microscopy

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