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. 2024:2694:133-150.
doi: 10.1007/978-1-0716-3377-9_7.

Single-Molecule Fluorescence Microscopy in Sensory Cilia of Living Caenorhabditis elegans

Elizaveta Loseva  1 , Jaap van Krugten  1 , Aniruddha Mitra  1 , Erwin J G Peterman  2
Affiliations

Single-Molecule Fluorescence Microscopy in Sensory Cilia of Living Caenorhabditis elegans

Elizaveta Loseva et al. Methods Mol Biol. 2024.

Abstract

Intracellular transport of organelles and biomolecules is vital for several cellular processes. Single-molecule fluorescence microscopy can illuminate molecular aspects of the dynamics of individual biomolecules that remain unresolved in ensemble experiments. For example, studying single-molecule trajectories of moving biomolecules can reveal motility properties such as velocity, diffusivity, location and duration of pauses, etc. We use single-molecule imaging to study the dynamics of microtubule-based motor proteins and their cargo in the primary cilia of living C. elegans. To this end, we employ standard fluorescent proteins, an epi-illuminated, widefield fluorescence microscope, and primarily open-source software. This chapter describes the setup we use, the preparation of samples, a protocol for single-molecule imaging in primary cilia of C. elegans, and data analysis.

Keywords: Caenorhabditis elegans; Live-cell imaging; Single-molecule imaging; Widefield fluorescence microscopy.

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References

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