Microfluidic-based imaging of complete Caenorhabditis elegans larval development.

Simon Berger,Alex Hajnal,Silvan Spiri,Andrew Demello

doi:10.1242/dev.199674

Abstract

ABSTRACTSeveral microfluidic-based methods for Caenorhabditis elegans imaging have recently been introduced. Existing methods either permit imaging across multiple larval stages without maintaining a stable worm orientation, or allow for very good immobilization but are only suitable for shorter experiments. Here, we present a novel microfluidic imaging method that allows parallel live-imaging across multiple larval stages, while maintaining worm orientation and identity over time. This is achieved through an array of microfluidic trap channels carefully tuned to maintain worms in a stable orientation, while allowing growth and molting to occur. Immobilization is supported by an active hydraulic valve, which presses worms onto the cover glass during image acquisition only. In this way, excellent quality images can be acquired with minimal impact on worm viability or developmental timing. The capabilities of the devices are demonstrated by observing the hypodermal seam and P-cell divisions and, for the first time, the entire process of vulval development from induction to the end of morphogenesis. Moreover, we demonstrate feasibility of on-chip RNAi by perturbing basement membrane breaching during anchor cell invasion.

Highlights

Immobilization of Caenorhabditis elegans larvae and adults during image acquisition is accomplished using agar pads, a simple construct consisting of a glass slide and a thin slab of agar onto which individual worms are placed (Luke et al, 2014; Goodman et al, 1998)
Device design and function Akin to the long-term immobilization approach proposed by Gokce et al (2017), we performed long-term imaging using a parallel array of trap channels, rather than multiple individual trap chambers
DISCUSSION we present a novel microfluidic long-term imaging platform that allows for high-resolution imaging and tracking of a variety of developmental processes in C. elegans during all four larval stages up to adulthood

Summary

Introduction

Immobilization of Caenorhabditis elegans larvae and adults during image acquisition is accomplished using agar pads, a simple construct consisting of a glass slide and a thin slab of agar onto which individual worms are placed (Luke et al, 2014; Goodman et al, 1998). As worms are immobilized in a secure fashion, a large variety of fluorescent markers and exposure times can be used Such an approach is flexible, minimizes any negative effects on the animal and allows the study of development across all larval stages. Parallelization is not achieved using multiple separate devices, but rather by using a number of parallel trap channels within a single device Such an approach significantly simplifies device setup and operation; repeatedly releasing worms from the trap channels into the large chamber does result in loss of animal identity as well as orientation. This in turn makes tracking developmental processes in the same worm impossible

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