Purchase this article with an account.
Kassandra Groux, Jules Scholler, Sacha Reichman, Michel Paques, Olivier Goureau, Jose Alain Sahel, Mathias Fink, Claude Boccara, Kate Grieve; Imaging retinal organoid subcellular dynamics non invasively with Full-Field OCT. Invest. Ophthalmol. Vis. Sci. 2019;60(9):3337.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
To image retinal organoids, as vehicles for progress in research on retinal diseases, non invasively at successive timepoints in their development, using Full-Field Optical Coherence Tomography (FFOCT).
Human retinal organoids generated from induced pluripotent stem cells at different stages of their development were used in our experiments. These structures were grown at the Vision Institute. We imaged twenty samples aged from 27 days to 126 days. We built two different setups to image the organoids: an FFOCT setup combined with multimodal fluorescence imaging, using known fluorescence labels, and an “inverted” FFOCT setup with an objective pointing upward in the sample arm to image from beneath, providing stability over long periods. The dynamic signal is created by intrinsic movements within cells, with a 1.5µm axial resolution and 0.5µm transverse resolution, up to 200µm depth.
The multimodal fluorescence imaging, used as a reference, allows us to distinguish different cells types, in both static and dynamic FFOCT images. We were able to distinguish dead cells, dying cells and living cells, each one creating different signals in static and dynamic FFOCT. Then, the dynamic imaging on the two setups revealed that the tracking of cells all day long and in real time is possible with our technique, allowing us to see e.g. division, death or differentiation of cells. Using our software, we are also able to reconstruct 3D images of organoids, allowing us to have a global vision of the intracellular dynamics of the organoid and the interactions between cells. Studying our computed images, we can see the difference in subcellular dynamics in retinal organoids of different ages, creating a new contrast mode for cells.
Using both static and dynamic FFOCT imaging creates a new valuable, label-free and non invasive contrast mode. We showed that static and dynamic FFOCT could potentially replace traditional imaging modalities for retinal organoids, without damaging them: we can image layers at different depths in a sample without using exogenous products as in fluorescence imaging, or without slicing as in histology. Our technique allows biologists to follow and control the evolution of the same organoid at different stages of its development.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
a) Timelapse movie (3 hours) of a 27 day-old retinal organoid, showing different cell behaviors b) 3D representation of the same sample
This PDF is available to Subscribers Only