Three-dimensional automated reporter quantification (3D-ARQ) technology enables quantitative screening in retinal organoids

Abstract

The advent of stem cell-derived retinal organoids has brought forth unprecedented opportunities for developmental and physiological studies, while presenting new therapeutic promise for retinal degenerative diseases. From a translational perspective, organoid systems provide exciting new prospects for drug discovery, offering the possibility to perform compound screening in a three-dimensional (3D) human tissue context that resembles the native histoarchitecture and to some extent recapitulates cellular interactions. However, inherent variability issues and a general lack of robust quantitative technologies for analyzing organoids on a large scale pose severe limitations for their use in translational applications. To address this need, we have developed a screening platform that enables accurate quantification of fluorescent reporters in complex human iPSC-derived retinal organoids. This platform incorporates a fluorescence microplate reader that allows -dimensional detection and fine-tuned wavelength selection. We have established optimal parameters for fluorescent reporter signal detection, devised methods to compensate for organoid size variability, evaluated performance and sensitivity parameters, and validated this technology for functional applications.

Large-scale Phenotypic Drug Screen Identifies Neuroprotectants in Zebrafish and Mouse Models of Retinitis Pigmentosa

“…To streamline such screens, we developed a high-throughput plate reader-based method for quantifying reporter gene expression in vivo ("ARQiv"; Walker et al, 2012). Recently, we adapted the ARQiv system to human stem cell-derived retinal organoids (Vergara et al, 2017) to enable cross-species PDD. To realize full throughput potential, ARQiv was combined with robotics-based automation to create "ARQiv-HTS" (G. White et al, 2016).…”

Organ‐on‐chip models: Implications in drug discovery and clinical applications

“…In an effort to improve quantitative analysis of organoid function for use in drug discovery, researchers have developed a screening platform that uses fluorophore technology to accurately and quickly assess organoid development and physiology (Vergara et al, ). The 3D automated reporter quantification (3D‐ARQ) utilizes a microplate reader to assess xyz‐dimensional detection and fine‐tuned wavelength selection (Vergara et al, ). The technology is quick, accurate, and reproducible, thus allowing it to be used in a number of other organoid systems in addition to the retina (Vergara et al, ).…”

The Emergence of Stem Cell‐Based Brain Organoids: Trends and Challenges

“…The pioneering work of the Sasai laboratory has uncovered these astonishing properties of pluripotent cells . Although culturing retinal organoids consumes substantial amount of time up to 150 days, several laboratories have successfully generated optic cups or retinal organoids harboring retinal pigment epithelia and functional photoreceptors …”