I am isolating gut cells for single cell RNA-sequencing and am currently measuring viability using 2 different methods: fixable viability dye Fluor 780 measured on the flow cytometer (this stains for amines in the cytoplasm of permeable [dead] cells) and MUSE Count and Viability reagent (this uses propidium iodide to bind DNA of dead cells). The two methods give drastically different results, with difference up to 40% in the assessment of live versus dead cells. Usually the MUSE detects more dead cells but the opposite occurs occasionally as well. Why might these huge discrepancies be happening and has anyone had any experience with comparing the different methods of detecting live versus dead cells and found how the results from different methods are similar/different? Which measurements are the most accurate?
IN my opinion you could use a simple Calcein AM viability assay. It is a scientifically accepted standard assay and it is well documented in various publications.
Hi Jayne, I agree with Andreas Eisenreich. Keep it simple and use tried and true methods like Calcein AM or trypan blue dye exclusion.
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