Lineage Tracing by Light-Sheet Microscopy and Computational Reconstruction.

Abstract

Lineage tracing based on modern live imaging approaches enables to visualize, reconstruct, and analyze the developmental history, fate, and dynamic behaviors of cells in vivo in a direct, comprehensive, and quantitative manner. Light-sheet fluorescence microscopy (LSFM) has greatly boosted lineage tracing efforts, because fluorescently labeled specimens can be imaged in their entirety, over long periods of time, with high spatiotemporal resolution and minimal photodamage. In addition, an increasing arsenal of commercial and open-source software solutions for cell and nuclei segmentation and tracking can be employed to convert data from pixel-based to object-based representations, and to reconstruct the lineages of cells in their native context as they organize in tissues, organs, and whole organisms. This chapter describes the preparation of LSFM image datasets and the use of three freely available platforms, namely, the Fiji/ImageJ plugins Massive Multiview Tracker (MaMuT), Mastodon and TrackMate, for small-scale and large-scale lineage tracing purposes using manual, semi-automated, and fully automated pipelines for nuclei or cell tracking. Lineage tracing with these tools is described on LSFM image datasets of fluorescently labeled embryos from the crustacean model Parhyale hawaiensis that lends itself to multi-scale investigations of development and regeneration.