Why You Need To Remove Dead Cellsĭead cells should be removed from all flow cytometry experiments that aim to evaluate live cell lineage and functionality.īelow are two simple examples of why you need to remove non-viable cells prior to implementing your flow cytometry gating strategies. These last two issues, in particular, will become even more troublesome if you are using dim markers or rare cell types. To make matters worse, both dead cells and apoptotic cells are highly autofluorescent. If you failed to remove your dead cells beforehand, gating on your population of interest will be exceptionally difficult. The problem here is that dead cells take up antibody very readily. In one final example, let’s say you’re simply staining surface antigens in a population of cells.
This means you will end up with fewer vital cells in one set of wells versus another, altering your results and negatively influencing your interpretation of the data. If you failed to remove your dead cells first, you could end up with different percentages of dead cells in one sorted sample versus another sample. Or maybe you’re sorting cells for a downstream functional assay, stimulating sorted T cells with antigen and measuring production of IFN. Since the dead cells in your sample will not divide, your culture will take extra time to reach the needed level of confluence, ruining your experimental timeline and weekend plans. If you fail to remove your dead cells first, you might think you’re seeding 10,000 cells, but in reality only 7,000 of your cells are actually viable. For this and other reasons, it’s important to remove dead cells from further analysis during your flow cytometry experiments.įor example, let’s say you merely need to generate an accurate cell count. This allows for antibodies to penetrate the cells, which can now mimic live cells. As cells die, the membrane becomes permeable.
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