Hello everyone!!!
I would like to analyze the desensitization and recycling of GPCRs following stimulation with their agonist. Have any of you ever performed such analysis? I would prefer to do it in fluorescence live imaging, but other suggestions are welcome. Would it be sufficient to use a fluorescent tag on my receptor to assess its inactivation (following internalization), or would it be preferable to use FRET or BRET probes to assess the response in terms of second messenger concentration or beta-arrestin engagement, respectively? And in the second case, are these sufficient parameters to claim that receptor signaling is inactivated?
I thank in advance anyone who answers.
Cornelius Krasel Thank you very much for your exhaustive answer.
So if I understand you correctly, to assess desensitization it would be sufficient for me to assess the change in cAMP concentration (via FRET) following agonist stimulation (we are talking about adrenergic receptors) and how this varies following subsequent stimulations over time? Whereas if I wanted to assess its trafficking, I could rely on a BRET assay. Am I right?
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