OrgaQuant: Human Intestinal Organoid Localization and Quantification Using Deep Convolutional Neural Networks

Abstract

Organoid cultures are proving to be powerful in vitro models that closely mimic the cellular constituents of their native tissue. Organoids are typically expanded and cultured in a 3D environment using either naturally derived or synthetic extracellular matrices. Assessing the morphology and growth characteristics of these cultures has been difficult due to the many imaging artifacts that accompany the corresponding images. Unlike single cell cultures, there are no reliable automated segmentation techniques that allow for the localization and quantification of organoids in their 3D culture environment. Here we describe OrgaQuant, a deep convolutional neural network implementation that can locate and quantify the size distribution of human intestinal organoids in brightfield images. OrgaQuant is an end-to-end trained neural network that requires no parameter tweaking; thus, it can be fully automated to analyze thousands of images with no user intervention. To develop OrgaQuant, we created a unique dataset of manually annotated human intestinal organoid images with bounding boxes and trained an object detection pipeline using TensorFlow. We have made the dataset, trained model and inference scripts publicly available along with detailed usage instructions.

Spatiotemporal Gradient and Instability of Wnt Induce Heterogeneous Growth and Differentiation of Human Intestinal Organoids

“…Various efforts have been made to overcome these challenges by increasing the number of experimental replicates, testing multiple lines, and validating the effect of different passage numbers to improve experimental statistics. Interestingly, organoids grown in a dome-shaped Matrigel scaffold have shown heterogeneity in size and shape, which has been reportedly observed in previous studies ( Kassis et al., 2019 ; Nanki et al., 2020 ). Yet, the underlying mechanism of this intra-heterogeneity in a Matrigel dome is unclear.…”

A Semi-automated Organoid Screening Method Demonstrates Epigenetic Control of Intestinal Epithelial Differentiation

“…Our toolkit, which we present in this article enables reproducible results across biological replicates using standard equipment and is suitable for screening setups. We established a quantification workflow that is based on the open source tools ImageJ and Ilastik, which is a simple yet robust alternative to recent stand-alone software options (Borten et al, 2018 ; Kassis et al, 2019 ) and could easily be adapted to different tissue organoids. In addition, qRT-PCR and flow cytometry of IEC lineages is sufficiently sensitive for initial screening and was subsequently confirmed by additional methods such as reporter organoids.…”

A semi-automated organoid screening method demonstrates epigenetic control of intestinal epithelial differentiation

“…Our toolkit, which we present in this article enables reproducible results across biological replicates using standard equipment and is suitable for screening setups. We established a quantification workflow that is based on the open source tools ImageJ and Ilastik, which is a simple yet robust alternative to recent stand-alone software options 46,47 and could easily be adapted to different tissue organoids. In addition, qRT-PCR and flow cytometry of IEC lineages is sufficiently sensitive for initial screening and was subsequently confirmed by additional methods such as reporter organoids.…”