Hi All,
I am planning to measure HIF expression with different cellular conditions using ELISA kit, from your experience what is the ideal number of cells used for this type of experiment?
Daer Reem!
Usually ELISA is performed in 96-well plates.
In this case, the number of cells depends on the size of the cells themselves, their metabolic activity and the volume of the well.
The larger the size of cells and their metabolism, the less they are needed per 1 well.
For example, for ELISA on human fibroblasts I used 25,000 cells per well, for C6, U251 gliomas - 10-12,000 per well.
Thanks for answering Dr Alexandr Chernov
I am using THP-1 differentiated macrophage, I will detach them using Trypl E then I will do cell lysis as I want to measure HIF expression. I think I need a high number of cells to have good HIF signal!
Is that true?
You can also check the recommended number of cells in the protocol provided by the producent of your ELISA kit. That may be a good suggestion.
And about your second question: high number of cells should give you high RNA concentration (if isolation is performed properly). Make sure that the concentration of your RNA is high enough to perform RT. If it's too low, you might consider using higher number of cells.
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