Hi all,
I have to do a phenotyping study on cell culture derived EVs,
in particular I should stain them using primary labelled ABs anti-EVs markers before proceeding to analyze them through high-sensitive flow cytometry and NTA in fluo-mode.
The issue is that the indicative "volume per test" reported in the ABs data-sheet is calibrated to defined number of cells in a determined volume,
how I could relate those values to EVs considering the different superficial areas and the different density of markers on their surface?
Hi Gabriele Vella
Sorry about this kind pf answer, but in my experience you just need/have to try. You need to standardize your own antibodies with your own EVs, because, as you said, different EVs have different markers with different densities. Besides, you need to consider antibodies clones and sellers.
In general, you could start with a dilution of 1:100 - 1:300 of your primary antibody. Maybe less if your antibodies are conjugated to a fluorophore.
Regards
Eduardo
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