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Improved methods for bulk cultivation and fixation of Loxodes ciliates for fluorescence microscopy

View ORCID ProfileBrandon Kwee Boon Seah, Christiane Emmerich, View ORCID ProfileAditi Singh, View ORCID ProfileEstienne Swart
doi: https://doi.org/10.1101/2022.05.29.493905
Brandon Kwee Boon Seah
Max Planck Institute for Biology, Max-Planck-Ring 5, 72076 Tübingen, Germany
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  • For correspondence: kb.seah@tuebingen.mpg.de
Christiane Emmerich
Max Planck Institute for Biology, Max-Planck-Ring 5, 72076 Tübingen, Germany
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Aditi Singh
Max Planck Institute for Biology, Max-Planck-Ring 5, 72076 Tübingen, Germany
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Estienne Swart
Max Planck Institute for Biology, Max-Planck-Ring 5, 72076 Tübingen, Germany
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  • ORCID record for Estienne Swart
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Abstract

Loxodes is one of the best ecologically characterized ciliate genera with numerous intriguing physiological abilities, including gravity-sensing organelles and nitrate respiration. However, these cells have been considered challenging to cultivate in bulk, and are poorly preserved by conventional fixatives used for fluorescence microscopy. Here we describe methods to grow and harvest Loxodes cells in bulk with liquid soil extract medium, as well as a new fixative called ZFAE (zinc sulfate, formaldehyde, acetic acid, ethanol) that can fix Loxodes cells more effectively than buffered formaldehyde or methanol. We show that ZFAE is compatible with immunofluorescence and the nuclear stain DAPI. Loxodes is thus now amenable to long-term maintenance, large-scale growth, and modern cell biology investigations of monoclonal strains in laboratory conditions.

Competing Interest Statement

The authors have declared no competing interest.

Footnotes

  • https://doi.org/10.17617/3.4ZQNBK

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY 4.0 International license.
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Posted May 29, 2022.
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Improved methods for bulk cultivation and fixation of Loxodes ciliates for fluorescence microscopy
Brandon Kwee Boon Seah, Christiane Emmerich, Aditi Singh, Estienne Swart
bioRxiv 2022.05.29.493905; doi: https://doi.org/10.1101/2022.05.29.493905
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Improved methods for bulk cultivation and fixation of Loxodes ciliates for fluorescence microscopy
Brandon Kwee Boon Seah, Christiane Emmerich, Aditi Singh, Estienne Swart
bioRxiv 2022.05.29.493905; doi: https://doi.org/10.1101/2022.05.29.493905

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