Looking for a live/dead assay, and not sure which one is best? Here we have the answer for you.
Why do a live/dead assay?
You should regularly track the viability of your cell culture, as it is representative of how healthy it is. The way you do this is by counting live and dead cells using a hemocytometer, and adding a dye that will selectively penetrate inside the dead (or apoptotic) cells, whose cellular membrane is damaged, while live cells will stay unstained. It’s really a simple method, and if you are counting cells with a hemocytometer anyway it’s worth to throw in a viability dye and count both populations (after all, most of us have two hands right? So don’t be stupid, count live with one counter and dead with the other one!).
What are the most commonly used dyes?
All viability dyes to be used together with a hemocytometer have the property of penetrating damaged cell membranes. The two most common are trypan blue and erythrosine B. Trypan blue is a blue (…) dye chemically derived from toluene. Erythrosin B is a cherry pink dye composed of a fluorescein salt (i.e. sensitive to light at certain wavelengths).
The duel: trypan blue vs. erythrosine BUnsure about which one fits your needs best? Have a look below.
|Trypan blue||Erythrosine B|
|Stains what?||Dead cells + serum proteins||Dead cells only|
|How much do I add?*||>400μg/mL||>5μg/mL|
|Time needed since cell death*||>50min||1min|
|In case you’re into fashion, what color?||blue||pink|
|In case you drop some, how bad is it?||Toxic (known carcinogenic)||It’s a food additive, what do you think? Bad for your clothes, maybe.|
|For cell lovers: how bad is it for the cells?||Toxic||Non toxic for periods of over 2h|
Overall, erythrosin B is a more reliable, less toxic method both for you and for the cells.