Multiplane HiLo microscopy with speckle illumination and non-local means denoising

Shuqi Zheng; Minoru Koyama; Jerome Mertz

doi:10.1117/1.JBO.28.11.116502

17 November 2023 Multiplane HiLo microscopy with speckle illumination and non-local means denoising

Shuqi Zheng, Minoru Koyama, Jerome Mertz

Author Affiliations +

Journal of Biomedical Optics, Vol. 28, Issue 11, 116502 (November 2023). https://doi.org/10.1117/1.JBO.28.11.116502

Abstract

Significance

HiLo microscopy synthesizes an optically sectioned image from two images, one obtained with uniform and another with patterned illumination, such as laser speckle. Speckle-based HiLo has the advantage of being robust to aberrations but is susceptible to residual speckle noise that is difficult to control. We present a computational method to reduce this residual noise without undermining resolution. In addition, we improve the versatility of HiLo microscopy by enabling simultaneous multiplane imaging (here nine planes).

Aim

Our goal is to perform fast, high-contrast, multiplane imaging with a conventional camera-based fluorescence microscope.

Approach

Multiplane HiLo imaging is achieved with the use of a single camera and z-splitter prism. Speckle noise reduction is based on the application of a non-local means (NLM) denoising method to perform ensemble averaging of speckle grains.

Results

We demonstrate the capabilities of multiplane HiLo with NLM denoising both with synthesized data and by imaging cardiac and brain activity in zebrafish larvae at 40 Hz frame rates.

Conclusions

Multiplane HiLo microscopy aided by NLM denoising provides a simple tool for fast optically sectioned volumetric imaging that can be of general utility for fluorescence imaging applications.

CC BY: © The Authors. Published by SPIE under a Creative Commons Attribution 4.0 International License. Distribution or reproduction of this work in whole or in part requires full attribution of the original publication, including its DOI.

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Shuqi Zheng, Minoru Koyama, and Jerome Mertz "Multiplane HiLo microscopy with speckle illumination and non-local means denoising," Journal of Biomedical Optics 28(11), 116502 (17 November 2023). https://doi.org/10.1117/1.JBO.28.11.116502

Received: 6 September 2023; Accepted: 3 November 2023; Published: 17 November 2023

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KEYWORDS

Speckle

Light sources and illumination

Denoising

Microscopy

Cameras

Fluorescence imaging

Tunable filters

Significance

Aim

Approach

Results

Conclusions

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