Next Article in Journal
A UV/Vis Spectroscopy-Based Assay for Monitoring of Transformations Between Nucleosides and Nucleobases
Next Article in Special Issue
Improving Signal and Photobleaching Characteristics of Temporal Focusing Microscopy with the Increase in Pulse Repetition Rate
Previous Article in Journal
Validity of Ultrasound Imaging Versus Magnetic Resonance Imaging for Measuring Anterior Thigh Muscle, Subcutaneous Fat, and Fascia Thickness
Previous Article in Special Issue
Enhanced Light Sheet Elastic Scattering Microscopy by Using a Supercontinuum Laser
Open AccessArticle

Highly Sensitive Shack–Hartmann Wavefront Sensor: Application to Non-Transparent Tissue Mimic Imaging with Adaptive Light-Sheet Fluorescence Microscopy

1
ITAV, Université de Toulouse, CNRS, 31106 Toulouse, France
2
Imagine Optic, 91400 Orsay, France
*
Author to whom correspondence should be addressed.
Methods Protoc. 2019, 2(3), 59; https://doi.org/10.3390/mps2030059
Received: 10 May 2019 / Revised: 25 June 2019 / Accepted: 8 July 2019 / Published: 11 July 2019
(This article belongs to the Special Issue Technical Advances in Light Microscopy)
  |  
PDF [2109 KB, uploaded 15 July 2019]
  |  

Abstract

High-quality in-depth imaging of three-dimensional samples remains a major challenge in modern microscopy. Selective plane illumination microscopy (SPIM) is a widely used technique that enables imaging of living tissues with subcellular resolution. However, scattering, absorption, and optical aberrations limit the depth at which useful imaging can be done. Adaptive optics (AOs) is a method capable of measuring and correcting aberrations in different kinds of fluorescence microscopes, thereby improving the performance of the optical system. We have incorporated a wavefront sensor adaptive optics scheme to SPIM (WAOSPIM) to correct aberrations induced by optically-thick samples, such as multi-cellular tumor spheroids (MCTS). Two-photon fluorescence provides us with a tool to produce a weak non-linear guide star (NGS) in any region of the field of view. The faintness of NGS; however, led us to develop a high-sensitivity Shack–Hartmann wavefront sensor (SHWS). This paper describes this newly developed SHWS and shows the correction capabilities of WAOSPIM using NGS in thick, inhomogeneous samples like MCTS. We report improvements of up to 79% for spatial frequencies corresponding to cellular and subcellular size features. View Full-Text
Keywords: light-sheet fluorescence microscopy; adaptive optics; highly-sensitive Shack–Hartmann; tissue mimics light-sheet fluorescence microscopy; adaptive optics; highly-sensitive Shack–Hartmann; tissue mimics
Figures

Figure 1

This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).
SciFeed

Share & Cite This Article

MDPI and ACS Style

Morgado Brajones, J.; Clouvel, G.; Dovillaire, G.; Levecq, X.; Lorenzo, C. Highly Sensitive Shack–Hartmann Wavefront Sensor: Application to Non-Transparent Tissue Mimic Imaging with Adaptive Light-Sheet Fluorescence Microscopy. Methods Protoc. 2019, 2, 59.

Show more citation formats Show less citations formats

Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Article Metrics

Article Access Statistics

1

Comments

[Return to top]
Methods Protoc. EISSN 2409-9279 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert
Back to Top