Jan 12, 2024
Preservation method for long-term storage of fluorescently labeled cells for microscopy
- Jacob Robertson1,
- Erin Garza2
- 1University of California San Diego;
- 2J. Craig Venter Institute
- JCVI West Protocols
Protocol Citation: Jacob Robertson, Erin Garza 2024. Preservation method for long-term storage of fluorescently labeled cells for microscopy. protocols.io https://dx.doi.org/10.17504/protocols.io.ewov1q15pgr2/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: December 19, 2023
Last Modified: January 12, 2024
Protocol Integer ID: 92469
Keywords: Preserve, Microscopy, Fluorescence, Long-term storage, Confocal, fluorescent microscope, labeled bacterial cell, cells for microscopy, preservation method, microscopy, detectable fluorescence, labeled cell, bacterial cell, fluorescence, cell integrity, cell, term storage
Abstract
This method can be used to preserve fluorescently labeled bacterial cells for long-term storage before imaging on a fluorescent microscope. Using this protocol, samples can be saved at 4°C for weeks-months while maintaining strong, easily detectable fluorescence and cell integrity.
Image Attribution
The image was produced by the authors using a Leica SP8 confocal microscope. This is a comparative photo of biofilms expressing a fluorescent protein that were preserved and imaged after 1 week vs 5 months.
Guidelines
This protocol has been successfully used on liquid bacterial cultures and on biofilms on solid plastic material. Cells were imaged using a confocal microscope up to 5 months after preservation. Fluorescence may be maintained longer than 5 months, but has not been tested by the authors.
Materials
Microfuge tubes
Glycerol
Paraformaldehyde
PBS
4ºC storage
Safety warnings
Paraformaldehyde is toxic and a skin irritant. Wear appropriate PPE when preparing and working with this solution.
Before start
Prepare the preservation components: 10% glycerol (sterile), 4% paraformaldehyde in PBS pH 7.4 (sterile)
Sample
2m
Collect your cells (up to 500 μl) in a sterile microfuge tube.
- This protocol can also be used on solid material containing biofilms.
2m
Preserve
3m
To your sample tube, add 250 µL sterile 10% glycerol and 250 µL sterile 4% paraformaldehyde in PBS (pH 7.4).
- If preserving solid material, make sure the entire sample is submerged in the preservation solution.
2m
Gently Mix
30s
Storage
1m
Store samples at 4 °C until ready to image.
1m