Protocol for fluorescent live-cell staining of tardigrades

Clayton J Harry; Jonathan D Hibshman; Amor Damatac 2nd; Phillip L Davidson; Martin Andres Estermann; Marycruz Flores-Flores; Caroline M Holmes; Jorge Lázaro; Elizabeth-Ann Legere; Jake Leyhr; Siddharthan Balachandar Thendral; Bridget A Vincent; Bob Goldstein

doi:10.1016/j.xpro.2024.103232

Protocol for fluorescent live-cell staining of tardigrades

STAR Protoc. 2024 Sep 20;5(3):103232. doi: 10.1016/j.xpro.2024.103232. Epub 2024 Aug 9.

Affiliations

¹ Department of Biology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
² Department of Biology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA; Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA.
³ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; Max Planck Institute for Evolutionary Biology, 24306 Ploen, Germany.
⁴ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; Department of Biology, Indiana University, Bloomington, IN 47405, USA.
⁵ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; Reproductive Developmental Biology Group, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, USA.
⁶ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; University of Colorado, School of Medicine, Anschutz Medical Campus, Department of Pediatrics, Section of Developmental Biology, 12801 E 17th Avenue, Aurora, CO 80045, USA.
⁷ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; Department of Organismal and Evolutionary Biology, Harvard University, Boston, MA, USA.
⁸ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; European Molecular Biology Laboratory (EMBL) Barcelona, Dr. Aiguader 88, 08003 Barcelona, Spain.
⁹ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; Department of Human Genetics, McGill University, Montreal, QC, Canada.
¹⁰ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; Department of Organismal Biology, Uppsala University, Uppsala, Sweden; Department of Biology, Duke University, Durham, NC 27708, USA.
¹¹ Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; Department of Biology, Duke University, Durham, NC 27708, USA.
¹² Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA; Department of Ecology, Evolution, and Marine Biology, University of California, Santa Barbara, Santa Barbara, CA 93106, USA.
¹³ Department of Biology, The University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA; Embryology Course, Marine Biological Laboratory, Woods Hole, MA 02543, USA. Electronic address: [email protected].

Abstract

Tardigrades are microscopic organisms with exceptional resilience to environmental extremes. Most protocols to visualize the internal anatomy of tardigrades rely on fixation, hampering our understanding of dynamic changes to organelles and other subcellular components. Here, we provide protocols for staining live tardigrade adults and other postembryonic stages, facilitating real-time visualization of structures including lipid droplets, mitochondria, lysosomes, and DNA.

Keywords: Cell Biology; Microscopy; Model Organisms.

MeSH terms

Animals
Lipid Droplets / metabolism
Lysosomes / metabolism
Microscopy, Fluorescence / methods
Mitochondria / metabolism
Staining and Labeling* / methods
Tardigrada* / cytology