Cheap and quick cell viability assay - (May/18/2006 )

Hi guys,

Does anybody know of a cell viability/vitality assay which is cheap and will not take decades to perform, can preferably be evaluated at a later time and most importantly not based on membrane permeability? (Or am I just dreaming)?

Thanks,
Casandra

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Look into MTT or MTS assay, although both are based on mitochondrial membrane permeability. Also check out the LDH release assay as another alternative.

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What about trypan blue? It has some membrane permability, but mostly its vitality by "pump ability". What cells are you looking at?

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You can try optical density using spectro?
You'll have to optimize your method. The cell samples should be of the same line/species. Determine the doubling time (DT) for your cell line. Then assume that if I have 50% and 100% viable cell (both of same OD, e.g. 0.1 to 0.3), after incubation within its DT, the OD will differ between the 50% and 100% viable cell samples.

That's troublesome too, initially, though. And you'll have to use other methods too, e.g. trypan blue exclusion or MTT/MTS assay (as suggested above) to verify.

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What do you consider cheap? How many viability assays are you going to be running?

I just used a kit that takes 10 minutes to determine viability via luminesence. It's about $75 per 200 50uL assays.

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Thanks guys but like I said I can't use membrane permeability, same goes for MTT, LDH and OD.
Haringsh, this luminescence assay is based on what? I'd appreciate it if you can give me the kit's name so I can it out. Ciao.
Casandra

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Hi Casandra,

The kit uses ATP from lysed cells to drive a luciferase reaction. The more viable cells you have, the more ATP you have, the more luminesence you have. Specifically, I used Promega's CellTiter-Glo Assay.

Hope that helps,
Hank

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trypan blue is virtually instant.

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if you have a cytometer, a simple FSC/SSC window would be sufficient to get accurate % of dead cells

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The Promega kit that I described goes from cells to results in 10 minutes. The problem with Trypan Blue is that you have to count the cells within 5 minutes of adding the stain, otherwise you'll get false positives. While this isn't a problem if you're working with one or two cell lines, trying to do anything high-throughput can prove extremely tedious.

Before trying the Promega assay I did TB counts for 4 different mutants in triplicate, but I could only stain and count one mutant each time without passing that 5 minute mark (I also do my counts in triplicate).

TB is definitely cheaper and easier if you're not doing anything in high volume. I suggest reading this paper if you're looking at an alternative to TB or some commercial kit:

http://www.ncbi.nlm.nih.gov/entrez/utils/l...sue=6&spage=693

I haven't had the opportunity to try this out yet, but it looks promising.

Hank

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