Hello, everyone
I’m currently working on a cycle cell analysis of HCT -116 using flow cytometry with a BD kit but I had a problem, I couldn’t run the sample because it had like a pellet which I can not solubilize.
I really hope you can help me with this problem
thank you.
Hi mónica,
how do you fix your cells? We use for that process icecold ethanol (70%) which can induce cell clumping but we resuspend on a vortex while we add the ethanol dropwise and just have a few doublets (or multiplets) of cells are formed. You can exclude multiplets on the cell cytometer by the following the protocoll of Wersto et al. (DOI: 10.1002/cyto.1171)
best wishes
You can aspirate the supernatant after pelleting and just add the PBS and vortex right before runing the sample. Even if your cells are moderately sticky, It will work fine.
Also If you do a lot of samples and run late to your flow appointment , it helps with time too.
Hi,
Try vortexing while fixing the cells. I use 70% ice-cold ethanol to fix cells. You could fix your cells by adding 70% ice-cold ethanol drop by drop with intermittent vortexing. If this step is not done properly, you are bound to get clumps which will persist further.
- Badges
- Science method