I am culturing mesenchymal stem cells on treated 1 mm^2 culture surfaces. This cannot be scaled up in area and may only be replicated in triplicate, i.e I have 3 mm^2 total area. If I assume that confluent cells are approx 625 µm^2 in area, then that will give me approx 1.5 x 10^3 cells per substrate. Is it best to pool these three samples to give me 4,500 cells total, or can I extract and detect proteins via microarray in triplicate with 1,500 cells? would DNA microarray be a better option?
It depends on the expression level of the proteins, quality of recognizing antibodies and the detection method. The abundance of high and moderate expressing proteins can be detected from the amount of cells that you have. Use infra-red fluorescence detection at 800 nm or another sensitive approach. If you have limited number of proteins it better to apply reverse phase protein arrays. If the number is much higher, antibody arrays are more appreciated (faster in a single assay). In the second case you cant apply small chamber supports (plane format of arrays) with low volume (50 microliters) and requiring 1-10 microgramme of total protein to be tested.DNA arrays might be more sensitive but you will answer about the abundance of transcripts and not of proteins. You should first test the method according to your goal.
Sorry, the correct sentence is
... If the number is much higher, antibody arrays are more appreciated (faster and in a single assay). In the second case you can apply small chamber supports (16-pad or format of plane arrays) requiring low volume (50 microliters) of 1-10 microgramme of total protein to be tested...
To close out this post, I can report that I abandoned protein microarray and focused instead on multiplex PCR. Specifically I employed the Fluidgm platform for analysis of nanolitre PCR samples.
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