FACS sorting affects cell function? - FACS sorting affects cell function (May/30/2007 )
I am new in Cytometry. I want to isolate CD8, CD4 using FACS sorting. I wonder if these T cells remain their function after staining with anti-CD4, CD8 for sorting.
In some study, I see that people use that sorting technique, but other studies use anti-CD4 to deplete CD4 cells in vivo.
Any comment would be appreciated.
I use cell sorting too but for other cell types. In my experience, cells do not change its behaviour or function. I use endothelial and smooth muscle cells. Maybe a critical issue is the survival rate of cells after cell sorting. Many cells dies after FACS, so I use a large number of cells to the FACS to recover cells with a number large enough to my assays. Manipulation is hard for cells, so take care of your cells after the FACS.
That technique about purifying a cell population adquiring the "contaminating cells" is also a good option. I use that for differentiated ES cells. If your interest population is bigger that contaminating populations, it could work.
With MACS also, you can deplete either one of the population you don't want or isolate either of the two you want. My senior used manual MACS to deplete CD4 population and for that I did the initial trial and it was good. I don't think it affected his results when he used the isolated CD8 for his experiments. But, may be it depends on for what you want T cells. Can you run a small trial and see if that is possible.