# Automated, high-throughput in situ hybridization of sea urchin (Lytechinus pictus) embryos

## TL;DR

This paper introduces an automated method to study gene activity in sea urchin embryos, allowing researchers to track 101 genes across early development stages efficiently.

## Contribution

The novel contribution is an automated high-throughput HCR pipeline for spatial gene expression profiling in sea urchin embryos.

## Key findings

- The HT-HCR pipeline processes 192 gene probe sets on embryos in 32 hours.
- High-quality localization data was obtained for 101 genes across three developmental stages.
- The method enables significant increases in throughput for spatial expression profiling in sea urchins.

## Abstract

Despite the reach of in situ hybridization (ISH) in developmental biology, it is rarely used at scale. The major bottleneck is the throughput of the assay, which relies upon labor-intensive manual steps. The goal of this study was to develop a high-throughput, automated hybridization chain reaction (HCR) pipeline for the sea urchin (Lytechinus pictus). Our method, which we term high-throughput (HT)-HCR, can process 192 gene probe sets on whole-mount embryos within 32 h. The physical properties of sea urchin embryos enabled us to utilize a 96-well plate format, miniaturized reaction volumes, a general-purpose robotic liquid handler and automated confocal microscopy. Using this approach, we produced high quality localization data for 101 target genes across three developmental stages. The results reveal the localization of previously undescribed physiological genes, as well as canonical developmental transcription factors. HT-HCR represents an order of magnitude increase in the throughput of spatial expression profiling studies utilizing the sea urchin. This will enable more-sophisticated perturbation analyses and drug-screening efforts in this emerging animal model.

Summary: An automated high-throughput HCR pipeline that enables high throughput gene expression profiling in sea urchin (Lytechinus pictus) embryos and the use of this tool to localize 101 genes in early development.