Abstract
Purpose :
Human retinal organoids (hRetOrg) are powerful emerging models to investigate human retinal development and disease. This project aims to develop a cellular-resolution visible-light optical coherence tomography (vis-OCT) prototype for their live morphological characterization.
Methods :
A vis-OCT system, which used light spectrum from 510 nm and 610 nm for imaging and operated at 50 kHz A-line sampling rate, was built at the University of Pittsburgh. hRetOrg at day (D) D43 (early-stage) and D113 (mid-stage) of development were maintained in culture medium and imaged live, noninvasively with vertical light beams focused by a scan lens (LSM03-VIS, Thorlabs). Each B-scan consisted of 500 A-lines with a scan density of 2-mm / line. Each volume had 500 B-scans and covered a 1×1-mm field of view. Scan acquisition was achieved by custom software written in LABVIEW and then further processed in MATLAB. After vis-OCT imaging, hRetOrgs were fixed, immunostained and examined in confocal and/or wide-field microscopy.
Results :
Live hRetOrg were successfully imaged at both developmental stages using the vis-OCT (Fig. 1). Their bodies and top and bottom surfaces can be appreciated by volumetric rendering. hRetOrgs were hollowed or semi-hollowed in the center. The neural retina was highly reflective, with retinal ganglion cells (RGC) in the innermost side of the organoid and photoreceptors (PR) in the outermost. At these developmental stages a thick pseudo-stratified layer was clearly detected in vis-OCT, and later confirmed by immunostaining in fixed hRetOrg. In addition, the retinal pigment epithelium (RPE) cells were concentrated adjacent to neural retina tissue. The mid-stage organoid, D113, was significantly larger, with a more obvious hollow and thicker neural retina when compared to the early-stage D43 one.
Conclusions :
Vis-OCT can provide 3-D, high-definition, and longitudinal imaging of live hRetOrg and might be a valuable tool for future organoid studies.
This abstract was presented at the 2023 ARVO Imaging in the Eye Conference, held in New Orleans, LA, April 21-22, 2023.