Dear experts,
Thanks for your insight into my previous queries.
could you please suggest what should be the appropriate cell number to get satisfactory results from cell lysate based ELISA? I am using MSC. Is T25 flask enoughfor the experiment, or can I use smaller plate?
Thanks,
priya
Hi Priya
It depends on the concentration of the protein you are interested in and also the affinity of the antibody. Some assays only require a single well of a 96 well plate while for others you may require more concentrated protein lysate.
Dr. Pool is correct. Every ELISA has a completely different sensitivity range and each cell line has a different range of that marker. Example for HUVECs, vimentin might be highly expressed on HUVEC cells and require very few cells, CD105 might be expressed in lower quantities and require more cells. Figure out your ELISA sensitivity, then look through the literature about your MSC cells to approximate the expression of your marker in these cells.
Hi Priya.
I agree with the previous comments, it will depend on your protein concentration and antibody affinity. In my opinion the best approach is start with an ELISA optimization, testing several concentrations of your protein in row, and several concentration of your antibody in column, for example. In addition, and based on my experience I usually coat the ELISA plate with 1.25-2.5 ug/ml of protein.
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