Hi, I need to quantify the levels of cAMP intracellularly in a cell line after stimulation of the EP2 receptor with PGE2. In the web I found severa protocols, most of them using radioactivity, which I can not use in my lab. I would appreciate any advice. Thanks Fidel
Hi. There are some FRET-based cAMP sensors utilizing EPAC as the sensing scaffold, which is flanked by fluorescent proteins (see this paper, for instance: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0122513 ). You will need a plasmid, a working transfection protocol and some sort of microplate reader, though.
You can also find kits for the detection of cAMP using antibodies. Kind of ELISA
You can use either FRET-based cAMP sensor as mentioned above or luciferase based assay. But these assays need special equipment. Instead you could try using cAMP assay kits available from companies such as Sigma and Abcam.
Thank you very much for the suggestions, I think I shall try the ELISA which is more affordable to me
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