Microscopy & Imaging JoVE (Open Access) Citable · DOI

Isotropic Light-Sheet Microscopy and Automated Cell Lineage Analyses to Catalogue Caenorhabditis elegans Embryogenesis with Subcellular Resolution

Name: Isotropic Light-Sheet Microscopy and Automated Cell Lineage Analyses to Catalogue Caenorhabditis elegans Embryogenesis with Subcellular Resolution
Uploaded: 2026-06-22
Duration: 8 min 16 s
Description: Caenorhabditis elegans (C. elegans) stands out as the only organism in which the challenge of understanding the cellular origins of an entire nervous ...

DOI: 10.3791/59533-v

What you'll learn

✓ Mount live C. elegans embryos for high-resolution light-sheet microscopy imaging
✓ Acquire isotropic volumetric data using dual-view inverted selective plane illumination microscopy
✓ Perform automated cell lineage tracing and identify individual cells with subcellular precision
✓ Quantify single-cell neurite outgrowth dynamics during embryonic neurogenesis

Protocol

Here, we present a combinatorial approach using high-resolution microscopy, computational tools, and single-cell labeling in living C. elegans embryos to understand single cell dynamics during neurodevelopment.

Difficulty

advanced

Total time

~4–6 hours per embryo (imaging + computational analysis)

Model organism

Caenorhabditis elegans

Biosafety

BSL-1

Steps

Prepare and mount C. elegans embryo samples

Harvest and position live C. elegans embryos on microscope slides with mounting medium to enable optical access for light-sheet imaging while maintaining embryonic viability.

▶ 00:34

Perform dual-view inverted selective plane illumination microscopy

Acquire high-resolution volumetric image stacks of living embryos using diSPIM from orthogonal viewing angles to generate isotropic 3D data with minimal phototoxicity.

▶ 01:48

Analyze cell lineage and segmentation computationally

Process volumetric microscopy data using automated computational tools to segment individual cells, assign cell identities, and reconstruct complete cell lineage trees with subcellular spatial resolution.

▶ 03:12

Quantify neurite outgrowth and cellular dynamics

Extract single-cell morphological parameters and track neurite extension trajectories over developmental time to characterize spatiotemporal patterns of neurogenesis.

▶ 06:06

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