|MadSci Network: Immunology|
Checking the cell viability is important for lots of reasons. Mainly because if you count 100 cells and only have 50% viability when you go to use them you may find that your experiment didn't work terribly well. Probably because you only had half the number of viable cells that you thought you did! In addition, there are certain things that can be recognised on dead cells and depending on how they died (necrosis or apoptosis ie the cell bursts or programmed cell death) and these could also affect the experiment. If I get your question correctly, you want to know why it's important to check the viability when using immunological techniques, I'd say that it's important for the above reason as well as the uptake of fluorochromes. When you phenotype cells by immunological methods eg flow cytometry, dead cells will take up the fluorochromes that are used to label the antibodies used in phenotyping. For example, if you wanted to phenotype CD4 and CD8 expression, dead cells may give false positives. They don't maintain membrane integrity and dyes and fluorochromes can enter into the cell making it red or green or both. The same would apply to other fluorescent techniques. Other dyes, such as propidium iodide (PI), can be used in flow cytometry to exlude dead cells. Dead cells allow PI to enter into the cells and the PI binds to DNA. These cells would appear red-ish by flow cytometry. So you could exlude all red-ish cells when phenotyping. On top of that, particularly if you wanted to phenotype cells using a flow cytometer, dead cells may have a different size and granularity from live cells. As flow cytometry also relies on looking at the size of cells, recognising dead cells is important. The cytometrist will probably be able to adjust the machine to exclude any dead cells (although this can be tough). Hope that answers your question.
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