Cultivation and Imaging of S. latissima Embryo Monolayered Cell Sheets Inside Microfluidic Devices.

Abstract

The culturing and investigation of individual marine specimens in lab environments is crucial to further our understanding of this highly complex ecosystem. However, the obtained results and their relevance are often limited by a lack of suitable experimental setups enabling controlled specimen growth in a natural environment while allowing for precise monitoring and in-depth observations. In this work, we explore the viability of a microfluidic device for the investigation of the growth of the alga Saccharina latissima to enable high-resolution imaging by confining the samples, which usually grow in 3D, to a single 2D plane. We evaluate the specimen's health based on various factors such as its growth rate, cell shape, and major developmental steps with regard to the device's operating parameters and flow conditions before demonstrating its compatibility with state-of-the-art microscopy imaging technologies such as the skeletonisation of the specimen through calcofluor white-based vital staining of its cell contours as well as the immunolocalisation of the specimen's cell wall. Furthermore, by making use of the on-chip characterisation capabilities, we investigate the influence of altered environmental illuminations on the embryonic development using blue and red light. Finally, live tracking of fluorescent microspheres deposited on the surface of the embryo permits the quantitative characterisation of growth at various locations of the organism.This work was supported by the CNRS, Sorbonne Université, the University of Cambridge, and, in part, by a grant from the Infinitus (China) Company Ltd (Contract Number RG82367) to G.S.K.S., by a career grant from the Swiss National Science Foundation (Grant Number P2EZP2_199843) to N.F.L., a project grant from the ARED Région Bretagne (Grant Number COH20020) to S.B., a CNRS grant (MITI “Lame Brune”) to B.C., and a project grant from the Gordon and Betty Moore Foun-dation (Symbiochip, Grant Number GBMF9333) to R.A.. The authors acknowledge open access funding by the Swiss National Science Foundation