What experiment can I perform to know how much time can my PYVMTs be without FBS and still be 99% viable?
I am currently working with PYVMTs and I need to know how much time can my cell line be fine (99% viable) in free serum media.
What can I do to know, preferably cheap.
Use trypan blue. Plate the cells at the concentration that you'd want to use them. If you don't know what concentration you'll want to use, try a few different ones to see how they look for you. Then harvest at set time points and count them. Should be pretty easy though a little time consuming to do manually. If you have an MTT or XTT type assay, you could always do that instead. A little more high-throughput and less time/labor intensive, though more expensive if you don't have the reagents for those assays.
No, I don't, I've never had to do that before, but should be pretty easy to set-up.
1. Plate 1000 cells (this is a random number but not unreasonable for a 96 well plate) in a 96 well plate. Use all the wells, so you have plenty for time points.
2. Harvest three wells per time point and count them. You know what your starting number is so you should have about that same number, give or take, at each time point. If you start to have die off, you'll see it over time. You need to have at least more than one well per time point as your counts will vary from well to well.
3. A reasonable starting set of time points could be 2h, 4h, 8h, 16h, 24h, 48h, etc. Then you'd just graph your counts and see where they start to fall off, if at all.
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