口回×oxygraphEilesHardwareCalibrateGraphToolsHelr足90Channel1APauseConfigureCalibrateStartStirrers300State1State 2MtrespiratoryrateMtrespiratory-fate-with-substratesNo:ADP..270withoutsubstrates线粒体240底物（琥珀酸盐，或著苹果酸盐+氨酸盐）State IADP210-tirisssssiiaaaaaaaoiiiiState II氧180气State 3aBAXO-Mt-reispiratory-rate-with-substrates+ADP150StateII12090Staté IV60State-4P/O=ADP/Oxygen consumedMtre'spiratoryratewhenADPirunout instate3300V1.5Time时间（分钟）Min20.0-75.00Oxygen3.0Rate
16 Assay of Mt Oxygen Consumption State 1 Mt respiratory rate without substrates State 2 Mt respiratory rate with substrates,No ADP State 3 Mt respiratory rate with substrates+ADP State 4 P/O=ADP/Oxygen consumed Mt respiratory rate when ADP run out in state 3
Oxidative phosphorylation ofmitochondriaState 1: Mt respiratoryrate without substratesState2: Mt respiratory rate with substrates, but noADPState 3: Mt respiratory rate with substrates + ADPState 4: Mt respiratory rate when ADP consumed in stare 3RCR (Respiratory Control Rate)RCR=State3/State4Intactness of mitochondria;functionof mitochondriaP/O ratio, P/O=amountofADP added/oxygen consumedEfficiencyofenergyconversion17
17 • State 1：Mt respiratory rate without substrates • State 2：Mt respiratory rate with substrates, but no ADP • State 3：Mt respiratory rate with substrates + ADP • State 4：Mt respiratory rate when ADP consumed in stare 3 • RCR (Respiratory Control Rate) RCR=State 3/State 4 Intactness of mitochondria; function of mitochondria • P/O ratio，P/O=amount of ADP added/oxygen consumed Efficiency of energy conversion Oxidative phosphorylation of mitochondria
ApplicationBiogerontology(2007)8:373-381ExampleDOI10.1007/s10522-007-9081-yRESEARCHARTICLED-Galactose toxicity in mice is associatedwith mitochondrial dysfunction: protecting effectsof mitochondrial nutrient R-alpha-lipoic acid***ControlB 2 x2ELAControlA3mD-GalLAD-Gal+LAmD-GalO2日D-Gal+LA1.5Fig.2RCRandADP/OofSuccinatelinkedmitochon-drialrespirationintheliverofC57BL/6mice.(A)RCR:(B)ADP/O.*P<0.05vs.Control,and**P<0.05vs.D-Gal18
18 Application Example
ApplicationBiogerontology(2007)8:373-381ExampleDO110.1007/s10522-007-9081-yRESEARCH ARTICLED-Galactose toxicity in mice is associatedwith mitochondrial dysfunction: protecting effectsof mitochondrial nutrient R-alpha-lipoic acidTable 1 Succinate linked mitochondrial respiration in thebrainofC57BL/6miceGroupState3State 4RespirationrespirationrespirationaftercccpaddedControl14.80± 1.2538.82 ± 4.9235.95±4.72LA36.57±6.6114.02± 4.0938.3 ± 3.42D-Gal29.70 ± 3.66*15.21± 4.6034.03±5.3136.22±3.00D-Gal + LA31.85±3.0013.20±2.24Abrain mitochondrial suspension was diluted with assaybufferto 1mg/ml and mitochondrialrespiration wasmeasured using oxygenelectrode.Respiration unitrepresented as nmol O2minmgprotein.Valuesaremean±SDoffiveanimals.P<0.05vs.control19
19 Application Example
YFHHIntroductionto theXFAnalyzerandtheXF Cell Mito Stress Test AssaySeahorseBioscience
O2 O2 O2 O2 O2 O2 H+ + H + H + H + H CO 2 CO2 CO2 CO2 H + CO2 O2 O2 O2 Introduction to the XF Analyzer and the XF Cell Mito Stress Test Assay