Are intracellular ionic concentrations accessible using fluorescent probes? The example of Mag-indo-1.
The study of the physicochemical properties of Mag-indo-1, a fluorescent probe used for intracellular magnesium measurements, has shown that in a biological environment the deprotonated form of this probe is in simultaneous equilibrium with a protonated form, a protein and a magnesium-bound form. The complex emission fluorescence spectrum emitted by a single living cell was analyzed using a computerized method, allowing the evaluation of the contribution of each species of the Mag-indo-1 to the cellular fluorescence. This approach used to evaluate intracellular Mg2+ concentration has also shown the variability of the important participation of protein-bound Mag-indo-1 to the cellular fluorescence. Thus the widely used ratioing method, unable to take into account this variability, cannot afford a reliable evaluation of [Mg2+]. Whatever the technique used for investigation (microfluorimetry, flow cytometry, etc.) the evaluation of [Mg2+]i using the fluorescent probe Mag-indo-1 requires a method able to quantify, in complex fluorescence, the fluorescence intensity of the forms involved in the equilibrium with Mg2+.
Morelle
B
B
Laboratoire de Chimie-Physique, Université de Perpignan, France.
Salmon
J M
JM
Vigo
J
J
Viallet
P
P
D016475:3T3 Cells; D000818:Animals; D003593:Cytoplasm / Q000737:chemistry / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007211:Indoles / Q000378:metabolism; D007424:Intracellular Fluid / Q000737:chemistry / Q000378:metabolism; D008274:Magnesium / Q000032:analysis* / Q000378:metabolism; D051379:Mice; D011485:Protein Binding; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods*
D005456:Fluorescent Dyes; D007211:Indoles; C088692:2-(4-(biscarboxymethyl)amino-3-(carboxymethoxy)phenyl)-1H-indole-6-carboxylic acid; D008274:Magnesium