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AAT Bioquest/Cell Meter™ JC-10 Mitochondrion Membrane Potential Assay Kit *Optimized for Microplate Assays*/22800/500
| Overview |  PrinterFriendlyVersion |
| Ex/Em(nm) | 510/525 |
| MW | N/A |
| CAS# | N/A |
| Solvent | N/A |
| Storage | F/D/L |
| Category | CellAnalysis CellCytotoxicity |
| Related | ApoptosisandCytotoxicity CellApoptosis BiochemicalAssays |
| Spectrum | AdvancedSpectrumViewer |
1.Preparecells:
1.1 Foradherentcells:Platecellsovernightingrowthmediumat20,000to80,000cells/well/90µLfora96-wellplateor5,000to20,000cells/well/20µLfora384-wellplate.
1.2 Fornon-adherentcells:CentrifugethecellsfromtheculturemediumandthensUSPendthecellpelletinculturemediumat100,000-200,000cells/well/90μLfora96-wellpoly-Dlysineplateor25,000-50,000cells/well/20μLfora384-wellpoly-Dlysineplate.Centrifugetheplateat800rpmfor2minuteswithbrakeoffpriortotheexperiments.
Note:Eachcelllineshouldbeevaluatedonanindividualbasistodeterminetheoptimalcelldensityforapoptosisinduction.
2.PrepareJC-10dye-loadingsolution:
2.1 Thawallthekitcomponentsatroomtemperaturebeforeuse.
2.2 Add50µLof100XJC-10(ComponentA)into5mLofAssayBufferA(ComponentB),andmixwell.
Note:Aliquotandstoretheunused100XJC-10(ComponentA)at-20oC.Avoidrepeatedfreeze/thawcycles.
3.RunJC-10assay:
3.1 Treatcellsbyadding10µLof10Xtestcompounds(96-wellplate)or5µLof5Xtestcompounds(384-plate)intothedesiredbuffer(suchasPBSorHHBS).
Note:Itisnotnecessarytowashcellsbeforeaddingcompound.However,iftestedcompoundsareserumsensitive,growthmediumandserumfactorscanbeaspiratedawaybeforeaddingcompounds.AddthesamevolumeofHHBSintothewells(suchas90µLfora96-wellplateor20µLfora384-wellplate)afteraspiration.Alternatively,cellscanbegrowninserum-freemedia.
3.2 Incubatethecellplateatroomtemperatureorina37oC,5%CO2incubatorforatleast15minutesoradesiredperiodoftime(forJurkatcells,4-6hourswithcamptothecinor3-5hourswithstaurosporinetreatment)toinduceapoptosis.
3.3 Add50µL/well(96-wellplate)or12.5µL/well(384-wellplate)ofJC-10dye-loadingsolution(fromstep2.2)intothecellplate(fromStep3.2).
3.4 Incubatethedye-loadingplateina37oC,5%CO2incubatorfor30-60minutes,protectedfromlight.
Note:Theappropriateincubationtimedependsontheindividualcelltypeandcellconcentrationused.Optimizetheincubationtimeforeachexperiment.
3.5 Add50µL/well(96-wellplate)or12.5µL/well(384-wellplate)ofAssayBufferB(ComponentC)intothedye-loadingplate(fromStep3.4)beforereadingthefluorescenceintensity.
Note1:DONOTwashthecellsafterloading.
Note2:Fornon-adherentcells,itisrecommendedtocentrifugecellplatesat800rpmfor2minuteswithbrakeoffafteraddingAssayBufferB(ComponentC).
3.6 MonitorthefluorescenceintensitiesatEx/Em=490/525nm(cutoffat515nm)and540/590nm(cutoffat570nm)withbottomreadmodeforratioanalysis.
| References&Citations |  CitationExplorer |
Below,youmayfindasmallsamplingofspecificJC-10™applications.ToinquireaboutapotentialapplicationofJC-10™,ortoconsultwithourfluorescentdyespecialists,pleasecontactusatsupport@AATbio.comor1-800-990-8053.
- High-contentassaysforhepatotoxicityusinginducedpluripotentstemcell–derivedcells.
ResearchersuseJC-10™tomonitormitochondrialdepolarizationasanindicationofhepatotoxicityandoxidativestressininducedpluripotentstemcell-derivedcells,withtheultimategoalofdesigningareliable,high-contentandimaging-basedinvitrotoxicityassay. - High-ContentHigh-ThroughputAssaysforCharacterizingtheViabilityandMorphologyofHumaniPSC-DerivedNeuronalCultures
JC-10™wasusedtostudyneuronsderivedfrominducedpluripotentstem-cells.SinceJC-10™willaccumulateinthemitochondriaofviablecells,itwasusedtodeterminecellviabilityinahigh-throughputassaycontext. - AnticancerActivityofNewSyntheticα-Methylene-δ-LactonesonTwoBreastCancerCellLines
ResearcherschoseJC-10™toinvestigatemitochondrialmembranepotentialandmembraneintegrityincellstreatedwithnaturalproducts,suchasα-Methylene-δ-Lactones,withthegoalofdevelopingnewtreatmentsforbreastcancer. - Tetrandrineprotectsmouseretinalganglioncellsfromischemicinjury.
JC-10™wasusedinflowcytometrytostudymitochondrialmembranepotential(ΔΨm)inprimaryculturedretinalganglioncells,asanextensionofthefieldofdrugdiscoveryintopreventionofischemicinjury. - Midazolaminducescellularapoptosisinhumancancercellsandinhibitstumorgrowthinxenograftmice
Inastudyofhumancancercells,JC-10™wasemployedtotrackcellularapoptosisasafunctionofmitochondrialmembranepotential,andconsequently,mitochondrialactivity.Researcherswereinterestedinthepossibleanestheticpropertiesofmidazolamforanticancerdrugdelivery. - MitochondrialproteomicswithsiRNAknockdowntorevealACAT1andMDH2inthedevelopmentofdoxorubicin-resistantuterinecancer
JC-10™wasusedbyresearchersforthepurposesofdrugdiscovery.Inparticular,researcherswereinterestedinfindingnewtreatmentsfordoxorubicin-resistantuterinecancer,usingJC-10™tomonitormitochondrialmembranepotentialandvalidatecellviabilityresults. - Coldexposurelowersenergyexpenditureatthecellularlevel
ResearchersusedJC-10™toinvestigatetherelationshipbetweentemperatureandcellularactivity.Inparticular,researcherswantedtoexploreifcoldtemperatureactsasastressoronmitochondrialmembranepotential,withregardstotheoxidativephosphorylationprocesswhichgeneratesATP. - Susceptibilityofgametesandembryosoftheeasternoyster,Crassostreavirginica,toKareniabrevisanditstoxins
JC-10™wasusedinthestudyofspermviability,fertilizationsuccesssandembryonicsurvivalofCrassostreavirginica.Specifically,JC-10™wasusedtoquantifymitochondrialmembranepotentialinspermcellsandtodeterminepossibletoxicityeffectsofalgalblooms. - CalmodulinantagoNISTsinducecellcyclearrestandapoptosisinvitroandinhibittumorgrowthinvivoinhumanmultiplemyeloma
JC-10™wasusedbyresearcherstostudycellcycleandapoptosisinhumanmultiplemyeloma.JC-10™functionedasaprobeforthedetectionofmitochondrialmembranepotentialdepolarization,whichwascrucialtothestudyofcaspaseactivatedapoptosis. - ActivationofthemitochondrialapoptoticpathwayproducesreactiveoxygenspeciesandoxidativedamageinhepatocytesthatcontributetolivertumOrigenesis
Researcherswereinterestedinthepathwaysinvolvedwithlivertumorigenesis.Tothatend,theyusedJC-10™tostudyactivationofapoptoticpathwaysrelatedtochangesinmitochondrialmembranepotential,withtheultimategoalofdiscoveringifantioxidanttherapymighthelpsuppresslivercarinogenesis.
