Hi everyone, I am conducting an experiment to induce p-PI3K activation using EGF. However, I am unsure about the appropriate volume of my medium + EGF mixture to add to each well of a 12-well plate. I plan to perform the assay at four time points: 1 min, 5 min, 10 min, and 15 min.
Thank you in advance for your advice..
Hi Maidoly Batista Fernández
In most cell culture protocols involving 12-well plates, the recommended volume of medium is typically 1 to 2 ml per well. This volume provides sufficient nutrients and space for the cells to proliferate and differentiate within the 3.5-3.8 cm² growth area of each well. 1 ml per well may be sufficient for your experiment, especially when it is a shorter one and if your cell line doesn’t have very high metabolic demands. Since EGF is present in the medium, its concentration and the cells' sensitivity to it will influence the optimal medium volume. Ensure the chosen volume maintains the desired EGF concentration and avoids potential depletion over time.
However, for longer assays or experiments where frequent media changes are not feasible, opting for a higher medium volume within the recommended range (1-2ml per well) can help sustain the cells for extended periods.
In general, when you must decide on the volume of media to be added per well, you will have to consider your specific experimental setup, cell type, and assay duration to provide ample room for growth, nutrient supply, and waste product dilution.
Best,
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