For my experiment, I want to grow bacterial cells in liquid medium under different conditions. One of the parameters I'm interested in is the cell numbers at the end of the treatment; I want to count CFUs as well as count cells using flow cytometry (syber9 staining) for this end.
I will need to preserve samples of the cultures for flow cytometry, and I am not sure if I should fix the cells in ethanol to store them in the medium term, or if I should freeze them at -20C. Which approach would be most appropriate? Thank you.
Ethanol fixation is an extremely straightforward and rapid procedure that allows cell structure to be conserved and is therefore suitable for flow cytometry, however it also results in a very restricted storage time and the introduction of certain artefacts that can interfere with your experiment.
freezing at -20°C resolves the issue and ensures long-term storage periods as well as minimal, if any, introduction of artefacts into the cells.
ensure that the laboratory in which you are working has suitable freezing equipment which you can access by rigorously implementing the freezing protocols.
Dear Miguel Angel Salinas Garcia
I completely agree with Souheib Khercha
If the staining you propose is based on the propidium iodide staining mechanism, SYTO 9 and SYBR Green I, you must take into account which of the two processes affects the integrity of the cell membrane more.
It is important to know which antigen your stained antibody labels in case the freezing protocol affects it.
You always have time to try both protocols.
Regards and good luck
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