Difference between revisions of "Magnesium Green"

Revision as of 10:19, 3 May 2019

high-resolution terminology - matching measurements at high-resolution

Magnesium Green

Description

Magnesium Green(MgG) belongs to the extrinsic fluorophores applied for measurement of mitochondrial ATP production with mitochondrial preparations. This fluorescent dye fluoresces when bound to Mg²⁺. The technique to measure mitochondrial ATP production is based on the fact that Mg²⁺ present different dissociation constants for ADP and ATP, and the adenine nucleotide translocase (ANT) exchanges ATP for ADP.

Abbreviation: MgG

Reference: Chinopoulos 2014 Methods Enzymol

MitoPedia methods: Fluorometry

Magnesium Green in high-resolution respirometry HRR

Instrument

O2k-FluoRespirometer with O2k-Fluo LED2-Module) selecting the Fluorescence-Sensor Blue/Filter Set MgG / CaG. Use black stoppers with black cover-slips to exclude disturbances by external light sources.

The fluorescent dye

Magnesium Green^TM is a registered trademark and available from Thermo Fisher Scientific (former: Invitrogen) in several formulations. For measuring mitochondrial ATP production a membrane impermeant formulation must be chosen (e.g. #M3733). Note that while its K_D for magnesium is 1.0 mM, its K_D for calcium is 6 µM: Magnesium Green binds stronger to calcium than to magnesium and cannot be used in the presence of significant concentrations of free calcium. Possible contaminations with transition metals should be complexed by a small (µM range) concentration of EDTA or DTPA.

Preparation of MgG solution

Experimental media for the MgG assay

K_d determination of ADP or ATP to Mg²⁺ using MgG

To assess the exchange of ADP/ATP by ANT using MgG, the K_d of ADP and ATP to Mg²⁺ should be previously calculated for the pertaining experimental conditions.

Useful information

» A kinetic assay of mitochondrial ADP-ATP exchange rate mediated by the adenine nucleotide translocase by [Chinopoulos C]

» Manual Fluorescent Magnesium Indicators.

@@ Line 13: / Line 13: @@
 == Magnesium Green in high-resolution respirometry [[HRR]] ==
 === Instrument ===
-[[O2k-Fluo LED2-Module]]) selecting the [[Fluorescence-Sensor Blue]]. Use [[Stopper\black PEEK\conical Shaft\central Port |black stoppers]] with [[Cover-Slip\black |black cover-slips]] to exclude disturbances by external light sources.
+::::* [[O2k-FluoRespirometer]] with [[O2k-Fluo LED2-Module]]) selecting the [[Fluorescence-Sensor Blue]]/[[Filter Set MgG / CaG]]. Use [[Stopper\black PEEK\conical Shaft\central Port |black stoppers]] with [[Cover-Slip\black |black cover-slips]] to exclude disturbances by external light sources.
-::::* [[O2k-FluoRespirometer]] with [[Fluorescence-Sensor Blue]]/[[Filter Set MgG / CaG]]
-::::* Magnesium Green and [[Calcium Green]] share the same fluorophore moiety in their molecular structure. Therefore, the optical properties are very similar and the same [[Fluorescence-Sensor Blue|fluorescence sensor]] and [[Filter Set MgG / CaG|filter set]] can be used.
-::::* Signal and output:
-::::::# Signal: The [[O2k-Fluo LED2-Module]] is operated through the  amperometric (Amp)-Channel of the O2k, with electric current (ampere  [A]) as the primary signal.
-::::::# Output: The focus of the output with Magnesium Green is on '''[[O2k signals and output| Type B]]''': Flow, flux, rate.
 === The fluorescent dye ===