Ask for advise for Growth Curve generation

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AllenChiu
AllenChiu's picture
Ask for advise for Growth Curve generation

I'm going to generate a growth curve with 4 slightly different A549 cell lines each infected with different mutant version of gene X.
I heard it's going to be complecated for counting.
And I do have some kind of Obsessive compulsive disorder when it comes to counting things.
Besides,I'm also warried about the dilution step which will add inaccuracy.
So I would like to know if I can turn to FACS for counting.
A question is should I count the whole amount of cell of each well or just a certain partition of it?
I think that the former will be more accurate since I'm not sure about if the cell will be Homogeneous during dilution.
Any advise from you is appreciated.
Thank you in advence!
PS:if I do have to count the cell number,should I use 24 well plate or 6 well ones?Is the cell amount per 24 well is enough for handling?

samm
samm's picture
Since you are working with an

Since you are working with an adherent cell line, you'll encounter losses as you pull 'em off the plate - which you'll need to do for flow/haemocytometer/Coulter counter and some absorbance methods. There are instruments that can count adherent cells on a plate, but they are in the $100k+ range, so unless you already have one, I won't recommend that!
An alternative is to use a soluble phase viable dye to track cell responses in real time over a series of time points (e.g redox potential) using a plate reader, or a cell division indicator such as CFSE/CPD670/DII (followed by flow cytometrric analysis - advantage being that you handle your cells much less).
For any of the above assays, 12 -24 well plates should work (100k cells+).

AllenChiu
AllenChiu's picture
thank you very much!

thank you very much!