Template:SUIT text D055: Difference between revisions

Latest revision as of 10:36, 8 June 2020

SUIT-006 MgG mt D055 is a protocol to investigate the O₂ flux and mitochondrial ATP production using the fluorescent dye Magnesium Green. In this protocol, the NADH Electron transfer-pathway state can be analyzed in mitochondrial preparations such as isolated mitochondria, tissue homogenates and permeabilized cells (already permeabilized when they are added to the chamber). Addition of the substrates pyruvate & malate (PM) to the mitochondrial preparation leads to LEAK state, and further addition of ADP induces the OXPHOS state, in which respiration is coupled to ATP production. After ADP addition, the signal from MgG gradually decreases, reflecting the ANT-mediated exchange of ADP for ATP, the later one having higher affinity for Mg²⁺. This exchange is stopped by addition of the ANT inhibitor carboxyatractyloside (Cat), which leads to LEAK state. With the addition of uncoupler, the Electron transfer pathway state can also be assessed in this protocol.

In the DatLab software, SUIT-006 MgG mt D055 DLP file is currently provided for the category N(PM). For using this protocol with other substrate/inhibitor combinations (e.g. succinate and rotenone for Succinate pathway control state or PMS for NS-pathway control state), a personalized DLPU can be created.

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-SUIT-006 MgG mt D055 is a protocol to investigate the O<sub>2</sub> flux and mitochondrial ATP production using the fluorescent dye [[Magnesium Green]]. In this protocol, the [[NADH Electron transfer-pathway state]] can be analysed in [[Mitochondrial preparations| mitochondrial preparations]] such as isolated mitochondria, tissue homogenates and permeabilized cells (already permeabilized when they are added to the chamber).
+SUIT-006 MgG mt D055 is a protocol to investigate the O<sub>2</sub> flux and mitochondrial ATP production using the fluorescent dye [[Magnesium Green]]. In this protocol, the [[NADH Electron transfer-pathway state]] can be analyzed in [[Mitochondrial preparations| mitochondrial preparations]] such as isolated mitochondria, tissue homogenates and permeabilized cells (already permeabilized when they are added to the chamber).
-Addition of the substrates [[pyruvate]] & [[malate]] (PM) to the mitochondrial preparation leads to [[LEAK-respiration|LEAK]] state, and further addition of ADP induces the [[OXPHOS]] state, in which respiration is coupled to ATP production. After ADP addition, the signal from MgG gradually decreases, reflecting the [[ANT]]-mediated exchange of [[ADP]] for [[ATP]], the later one having higher affinity for Mg<sup>2+</sup>. This exchange is stopped by addition of the ANT inhibitor [[carboxyatractyloside]] (Cat), which leads to [[LEAK-respiration|LEAK]] state. With the addition of [[uncoupler]], the [[ET]] state can also be assessed in this protocol.
+Addition of the substrates [[pyruvate]] & [[malate]] (PM) to the mitochondrial preparation leads to [[LEAK respiration|LEAK]] state, and further addition of ADP induces the [[OXPHOS]] state, in which respiration is coupled to ATP production. After ADP addition, the signal from MgG gradually decreases, reflecting the [[ANT]]-mediated exchange of [[ADP]] for [[ATP]], the later one having higher affinity for Mg<sup>2+</sup>. This exchange is stopped by addition of the ANT inhibitor [[carboxyatractyloside]] (Cat), which leads to [[LEAK respiration|LEAK]] state. With the addition of [[uncoupler]], the [[Electron transfer pathway]] state can also be assessed in this protocol.
+In the DatLab software, SUIT-006 MgG mt D055 DLP file is currently provided for the category N(PM). For using this protocol with other substrate/inhibitor combinations (e.g. [[succinate]] and [[rotenone]] for [[Succinate pathway control state]] or [[PMS]] for [[NS-pathway control state]]), a personalized [[Export DL-Protocol User (*.DLPU)|DLPU]] can be created.