Dear all, I am using confocal leaser microscope for calcium imaging. Last couple of days I have faced difficulty to get the cell responses, even though I did not get the response of ATP. I am using HEK293T cells and transfect plasmid DNA using ScreenFect (Wako). I would like to see the cell responses of different agonists and mixtures of known agonists and putative antagonist. I have always used 5 µM ATP for normalizing cell activity. But, last few days I did not get any response from ATP. Can you please suggest me about this problem.
Thank you in advance.
Dear Bapon,
There are several possibilities why you are not seeing any ATP responses. Your ATP is infected by bacteria and chewed up. Or your transfection did not work and your cells are not expressing any ATP receptors. Or your cells are are depolarised and resting membrane potential close 0 mV and therefore the cells are not responding to ATP. I have done expression in oocytes before and studied ATP-gated ion channels. See my publications listed below.
best wishes, Refik
Article Allosteric Modulation of Native Cochlear P2X Receptors: Insi...
Thesis Molecular physiology of ATP-gated ion channels assembled fro...
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