In case microbial samples, e.g. environmental water samples need to be stored or transported for flow cytometry analysis, what would be a good method for preserving integrity of cell membranes? Main goal is to do live & dead cell count with SYBRGreen and PI dyes.
Would freezing in glycerol be ok? I was using it for transfected cells but not sure it would work here. Would be grateful for any input.
For 1-2 days it could be stored at around 4-8 degrees C until analysis as even freezing them in cryoprotectants can kill a few cells. Would suggest freezing only if it is for a long term storage and since your aim is live dead analysis I would not suggest storing it.
Hello, when I did flow cytometry I kept the samples at 4°C before live-dead analysis, but always passed them to cytometry on the same day as collection. I never froze the samples so do not know for the freezing, it could disrupt the membranes and lead to increased number of dead cells.
Dear Ashutosh Acharya and Lisa Zoé Auclert, thank you so much for your answers. Let me provide some background. The samples need to be frozen in order to be shipped for flow cytometry analysis. I have now tried the glycerol solution and it seems to be working well enough, although I have the feeling it is producing artifact counts, maybe the viscosity of the sample or a chemical effect. I am still investigating this.
The live cells detection with the double staining seems to work reasonably well. However, the single staining with SYBRGreen for total cell detection seems to produce a lot of artifact counts and non reproducible results. Do you move the gate around to select the correct population? The protocol I follow suggests to keep the gate as is after defining it. Or so you have any other suggestions why this is happening?
- Badges
- Science topic
- Similar topics
- Biological Science
- Biochemistry
- Enzymology
- Enzymes