Made a mistake and fixed cells with 30% ethanol for cell cycle analysis. Can I s - (Oct/02/2014 )
I was fixing mammalian cells for propidium iodide staining then cell cycle analysis. However, I made a careless mistake and fixed my cells with 30% ethanol instead of 70% ethanol. I realized my mistake right after fixing one set of cells. But a second set of cells were fixed with 30% ethanol then sat at -20C for about 30 minutes. I added ethanol to both sets of cells to put them in a 70% solution. Cells were vortexed while adding this additional ethanol.
Two set of cells - details
1. Fixed cells with 30% ethanol. Realized the mistake right after adding the 30% ethanol. Immediately added ethanol to make it a 70% solution.
2. Fixed Cells with 30% ethanol and placed in -20C for 30min. I removed these cells from -20C and added ethanol to make it a 70% solution.
Note: I vortexed these cells lightly when adding the ethanol (i.e. 30% up to a 70% solution). Both sets of cells are now stored at -20C.
Can I still stain these cells and perform cell cycle analysis? What is the consequence of these cells being in 30% ethanol for short period of time then switching to a 70% ethanol solution? Cell clumping, fragmenting, distorted shape/size? I performed treatment on these cells that took a while so I would like to salvage these cells if possible.
Thanks!
No one replied to this post so I will provide some feedback. I finally got around to processing these samples and the cell cycle analysis worked even though the cells were not fixed in a traditional manner. The cells were stored at -20C for 3 days.