June 2020
Volume 61, Issue 7
Free
ARVO Annual Meeting Abstract  |   June 2020
Optical phase contrast imaging of human retinal cells by changing the tissue refractive index
Author Affiliations & Notes
  • Laura Kowalczuk
    Laboratory of Applied Photonic Devices, Ecole Polytechnique Federale de Lausanne, Lausanne, Switzerland
    Department of Ophthalmology, University of Lausanne, Switzerland
  • Timothe Laforest
    Laboratory of Applied Photonic Devices, Ecole Polytechnique Federale de Lausanne, Lausanne, Switzerland
    EarlySight SA, Switzerland
  • Mathieu Künzi
    EarlySight SA, Switzerland
  • Florentino Caetano dos Santos
    Laboratory of Applied Photonic Devices, Ecole Polytechnique Federale de Lausanne, Lausanne, Switzerland
  • Alexandre P Moulin
    Department of Ophthalmology, University of Lausanne, Switzerland
    Jules-Gonin Eye Hospital, Fondation Asile des aveugles, Switzerland
  • Leonidas Zografos
    Department of Ophthalmology, University of Lausanne, Switzerland
    Jules-Gonin Eye Hospital, Fondation Asile des aveugles, Switzerland
  • Patricia Lassiaz
    UMR_S 1138, Team 17, INSERM, France
    Centre de Recherche des Cordeliers, University of Pierre et Marie Curie, Paris Descartes University, Sorbonne Paris Cité, France
  • Francine F Behar-Cohen
    UMR_S 1138, Team 17, INSERM, France
    Department of Ophthalmology, Ophthalmopole, Cochin Hospital, Assistance Publique - Hôpitaux de Paris, France
  • Christophe Moser
    Laboratory of Applied Photonic Devices, Ecole Polytechnique Federale de Lausanne, Lausanne, Switzerland
  • Footnotes
    Commercial Relationships   Laura Kowalczuk, None; Timothe Laforest, Earlysight (I); Mathieu Künzi, Earlysight (F); Florentino Caetano dos Santos, None; Alexandre Moulin, None; Leonidas Zografos, None; Patricia Lassiaz, None; Francine Behar-Cohen, Earlysight (I); Christophe Moser, Earlysight (I)
  • Footnotes
    Support  EIT Health ASSESS project
Investigative Ophthalmology & Visual Science June 2020, Vol.61, 1854. doi:
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      Laura Kowalczuk, Timothe Laforest, Mathieu Künzi, Florentino Caetano dos Santos, Alexandre P Moulin, Leonidas Zografos, Patricia Lassiaz, Francine F Behar-Cohen, Christophe Moser; Optical phase contrast imaging of human retinal cells by changing the tissue refractive index. Invest. Ophthalmol. Vis. Sci. 2020;61(7):1854.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Based on oblique partially coherent illumination of transparent samples, we developed a simple custom Optical Phase Imaging (OPI) microscope providing a label-free, semi-quantitative phase contrast imaging. The aim of this study was to explore this ex-vivo modality for retinal imaging and correlate it with standard clinical images and fluorescence microscopy.

Methods : Multimodal macular imaging was performed on the flat-mounted retina of an eye presenting an epiretinal membrane with cystoid macular edema, enucleated for a peripheral melanoma. After glial fibrillary acidic protein (GFAP) - aquaporin (AQP)-4 – collagen (Col)-IV co-immuno-labeling and nuclei staining, the retina was cleared by index matching in a medium of refractive index (RI) 1.46 to decrease scattering for high-resolution deep-tissue ex vivo imaging. We performed a comparison of the clinical examinations obtained by Optical Coherence Tomography-Angiography and fluorescein angiography before enucleation, with the images obtained with confocal microscopy and OPI microscopy. Ex-vivo imaging of the retina mounted in a medium with a lower RI (1.40), close to the mean RI of Muller glial cell (MGC), was then repeated to better view the latter cells.

Results : The retinal vessels were used as landmarks for correlating all imaging modalities. OPI microscopy allowed for different contrast imaging depending on the RI of the mounting medium. With the high RI medium (1.46), deep contrast imaging of nuclei and intraretinal cysts was obtained. The solution with a RI of 1.4 provided an improvement in the contrast of the retinal structures, from the inner layer (AQP4-positive MGC, epi-retinal membrane, nerve fibers surrounded by GFAP-positive astrocytes) to the photoreceptor segments. No AQP4 labeling was observed inside the cyst. AQP4-positive, GFAP-negative cells were visualized on the ColIV-labeled epi-retinal membrane, demonstrating that the membrane is made of retinal Muller glial cells.

Conclusions : This morphological correlative imaging study demonstrated OPI on numerous cellular structures of a human retina by tuning the tissue RI. This label-free in-depth imaging modality offers a new research tool to study the cellular origin of retinal diseases.

This is a 2020 ARVO Annual Meeting abstract.

 

Comparison of clinical (A), fluorescence (B) and phase contrast (C-E) images, in nasal (1), central (2) and temporal (3) macula and in temporal periphery (4) of a human retina.

Comparison of clinical (A), fluorescence (B) and phase contrast (C-E) images, in nasal (1), central (2) and temporal (3) macula and in temporal periphery (4) of a human retina.

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