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When setting up a cell sorting experiment, there are many things to consider.

You must consider which controls you’re going to use, how you’re going to compensate the experiment, which instrument and which instrument settings are ideal, and how you plan to analyze, gate, and present your data. With so many things to consider, it’s easy to lose site of the small things that can drastically affect the viability of your cells, including the composition of your suspension buffer. The composition of the suspension buffer for preparation, staining, analyzing and sorting is perhaps the most important parameter for maintaining viability during a cell sorting experiment. While the precise components of a buffer can differ depending on the cell type, there are a 5 key points to keep in mind.

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There are several methods for analyzing live, dead, and apoptotic cells by flow cytometry.

As cells die, the membrane becomes permeable. This allows for antibodies to penetrate the cells, which can now mimic live cells. For this and other reasons, it’s important to remove dead cells from further analysis during your flow cytometry experiments. For example, let’s say you merely need to generate an accurate cell count. 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. Since the dead cells in your sample will not divide, your culture will take extra time to reach the needed level of confluence. Don’t make the mistake of forgetting to add a live-dead cell marker to your next flow cytometry experiment. Here are the top 3 markers available to you.

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