I want to check calcium signalling in cells by Florescence imaging.
The best way, I think, is to exchange Calcium ions by Sodium ions with a cationic exchange resin. Then, its much easier to get free from the sodium ions. Or maybe, the Sodium ions do not interfere with your experiment
Hello Saravanakumar,
I little more information might be useful, but off the top of my head I would think of EDTA or EGTA to complex free Ca-ions. It depends on the experimental setup. We did calcium signalling in cells without bothering much of the exterior Calcium level. If you provide more details it might be easier to get good advice.
Cheers
Sebastian
Are you looking at fixed or live cells by IFA?
The reason I ask is that placing live cells into a Ca-free environment will cause intracellular Ca to be released from intracellular stores (ER, calcineurin, calmodulin, etc) in attempt to provide Ca to essential survival pathways - it is a cofactor for many critical enzymes. This is likely to adversely effect what is is you are trying to measure.
If this ins't a concern, then using ion chelators like EDTA and EGTA are very useful. EDTA(IV) binds to a metal cations through its two amines and four carboxylates, forming a ). EDTA targets most divalent cations (EDTA forms especially strong complexes with Mn(II), Cu(II), Fe(III), Pb (II) and Co(III); it also captures Ca(II) and Mg(II)) while EGTA is more selective for Ca.
I recommend looking at Wikipedia for details on mechanism !
hi thanks , I want to check calcium ion signalling in RBC cells usinfg fluor 4
EDTA or EGTA will help to chelate any Ca2+ leak for the Ca2+ - Na+ exchanger from the cells. This will be much useful if the buffer itself is Calcium free from onset.
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