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Millipore/AG325 | Fluoro-Jade® C/AG325/50 mg
  • Millipore/AG325 | Fluoro-Jade® C/AG325/50 mg

Millipore/AG325 | Fluoro-Jade® C/AG325/50 mg

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货号: AG325
品牌: Millipore
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    • Description
      CatalogueNumberAG325
      BrandFamilyChemicon®
      TradeName
      • Fluoro-Jade
      • Chemicon
      DescriptionFluoro-Jade®C
      OverviewFluoro-Jade®C,likeitspredecessors,Fluoro-Jade®andFluoro-Jade®B,werefoundtostainalldegeneratingneurons,regardlessofspecificinsultormechanismofcelldeath.Therefore,thepatternsofneuronaldegenerationseenfollowingexposuretoeithertheglutamateagoNIST,kainicacid,ortheinhibitorofmitochondrialrespiration,3-NPA,werethesameforalloftheFluoro-Jadeâdyes.However,therewasaqualitativedifferenceinthestainingcharacteristicsofthethreeFluorochromes.Specifically,Fluoro-Jade®Cexhibitedthegreatestsignaltobackgroundratio,aswellasthehighestresolution.Thistranslatestoastainofmaximalcontrastandaffinityfordegeneratingneurons.Thismakesitidealforlocalizingnotonlydegeneratingnervecellbodies,butalsodistaldendrites,axonsandterminals.ThedyeishighlyresistanttofADIngandiscompatIBLewithvirtuallyallhistologicalprocessingandstainingprotocols.TriplelabelingcanbeaccomplishedbystainingdegeneratingneuronswithFluoro-Jade®C,cellnucleiwithDAPIandactivatedastrocyteswithGFAPimmunofluorescence.



      APPEARANCE:Coffeebrowntobrickredpowder.

      MOLECULARWEIGHT:823

      EXCITATIONPEAK:485nm

      EMISSIONPEAK:525nm

      FILTERSYSTEM:Fluorescein/FITC

      SOLUBILITY:Highlysolubleinwaterandbases,moderatelysolubleinalcoholandweakacids.

      TOXICITY:Althoughthecompoundappearstobeoflowtoxicity,ithasnotbeenextensivelyevaluatedandthereforeroutinelaboratorycautionshouldbeexercised.Notintendedforhumanconsumption.
      BackgroundInformationFluoro-JadeC,likeitspredecessors,Fluoro-JadeandFluoro-JadeB,werefoundtostainalldegeneratingneurons,regardlessofspecificinsultormechanismofcelldeath.Therefore,thepatternsofneuronaldegenerationseenfollowingexposuretoeithertheglutamateagonist,kainicacid,ortheinhibitorofmitochondrialrespiration,3-NPA,werethesameforalloftheFluoro-Jadedyes.However,therewasaqualitativedifferenceinthestainingcharacteristicsofthethreefluorochromes.Specifically,Fluoro-JadeCexhibitedthegreatestsignaltobackgroundratio,aswellasthehighestresolution.Thistranslatestoastainofmaximalcontrastandaffinityfordegeneratingneurons.Thismakesitidealforlocalizingnotonlydegeneratingnervecellbodies,butalsodistaldendrites,axonsandterminals.Thedyeishighlyresistanttofadingandiscompatiblewithvirtuallyallhistologicalprocessingandstainingprotocols.TriplelabelingcanbeaccomplishedbystainingdegeneratingneuronswithFluoro-JadeC,cellnucleiwithDAPIandactivatedastrocyteswithGFAPimmunofluorescence.Fluoro-Jade™isaregisteredtrademarkofHisto-Chem,Inc.
      ProductInformation
      StorageandShippingInformation
      StorageConditionsThelyophilizedpowdershouldbestoredwellsealedatroomtemperature,preferableinadesiccator(duetoitshygroscopicnature)foruptooneyearfromdateofreceipt.Theliquidstocksolution(0.01%)indistilledwatercanbestoredat2-8°Cforupto3months.The.0002-.0001%workingsolutionin0.1%aceticacidshouldbeusedwithin4hrsofpreparation.
      Applications
      KeyApplications
      • Immunohistochemistry
      ApplicationNotesSUGGESTEDPROTOCOLFORUSINGFLUORO-JADE®C

      Processing:Halfofeachgroupofbrainswereparaffinembeddedandcutonarotarymicrotomewhiletheremainderwerecutonafreezingslidingmicrotome.Paraffinsectionswere10uminthicknesswhilefrozensectionswerecutatathicknessof25um.Priortostaining,sectionsweremountedfromdistilledwaterontogelledslides.Gelatincoatedslideswerepreparedbyimmersionina60degreeCsolutionof1%pigskingelatin(Sigma;typeA,300Bloom)andthenovendriedovernightatthesametemperature.Thesectionsweremountedontotheslidesfromdistilledwaterandthenairdriedforatleast30minonaslidewarmerat50degreesC.Slidesbearingfrozencuttissuesectionswerefirstimmersedinabasicalcoholsolutionconsistingof1%sodiumhydroxidein80%ethanolfor5min.Theywerethenrinsedfor2minin70%ethanol,for2minindistilledwater,andthenincubatedin0.06%potassiumpermanganatesolutionfor10min.Followinga1-2minwaterrinse,theslideswerethentransferredfor10mintoa0.0001%solutionofFluoro-Jade®Cdissolvedin0.1%aceticacidvehicle.Theproperdilutionwasaccomplishedbyfirstmakinga0.01%stocksolutionofthedyeindistilledwaterandthenadding1mLofthestocksolutionto99mLof0.1%aceticacidvehicle.Theworkingsolutionwasusedwithin2hofpreparation.Thestocksolution,whenrefrigerated,canbekeptforlongperiodsbutshouldbediscardedifthesolutionbecomescloudy.Theslideswerethenrinsedthroughthreechangesofdistilledwaterfor1minperchange.Excesswaterwasdrainedontoapapertowel,andtheslideswerethenairdriedonaslidewarmerat50degreesCforatleast5min.Theairdriedslideswerethenclearedinxyleneforatleast1minandthencoverslippedwithDPX(FlukaorSigma)nonfluorescentmountingmedia.Polarcoverslippingmedia,suchasthosethatcontainwater,alcoholorglycerolwereneverused.Forcomparativepurposes,someslideswerestainedwithFluoro-Jade®Baccordingtothepreviouslydescribedprocedure.Whenworkingwithparaffinprocessedtissue,thesectionsarefirstdeparaffinizedthroughtwo10minchangesofxyleneandthenthesectionsarerehydratedthroughagraduatedalcoholseries,omittingthebasicalcoholsolution.Onceindistilledwater,thesectionsaretransferredtothepotassiumpermanganatesolutionatwhichpointthestainingprocedureisidenticaltothatdescribedforfrozensections.Multiplelabeling:Fluoro-Jade®CcanreadilybecombinedwithotherfluorescentMarkers.Multiplelabelingwasachievedusinganti-glialfibrillaryacidicprotein(GFAP)immunocytochemistrytolabelactivatedastrocyteswhileusingDAPItolabelnuclearDNA.Incorporating4",6-diamidino-2-phenylindole(DAPI;Sigma,St.LouisMO)asafluorescentnuclearstainisaccomplishedbysimplyincorporating0.0001%intotheFluoro-Jade®Cstainingsolution.Thisisaccomplishedbytheadditionof1mlof0.01%DAPIstocksolutionto99mlof0.1%aceticacid.Fluoro-Jade®CwasalsocombinedwithimmunofluorescentlabelingofGFAPaccordingtothefollowingprocedure.Loosefrozentissuesectionswereincubatedinapredilutedsolutionofanti-GFAP(ChemiconhasanumberofdifferentantibodiestoGFAP)atabout5degreesCintherefrigeratorfor1-3days.Itshouldbementionedthatalthoughinthisstudyallimmunocytochemistrywasperformedonfrozensections,themethodsarefullycompatiblewithparaffinprocessedtissueaswell.Sectionswererinsedintwochangesofbufferedsalinefor10mineachandthentransferredtoatetramethylrhodamineisothiocynate(TRITC)labeledsecondaryantibody(ChemiconhasanumberofdifferentTRITClabeledsecondaryantibodies),diluted1:100inbufferedsaline,for1hatroomtemperature.Sectionswererinsedintwochangesofbufferedsalinefor10mineachandthenthesectionsweremountedontogelledslidesfromdistilledwaterandairdriedonaslidewarmerat50degreesCfor30min.TocombinewithFluoro-Jade®C,theslidemountedsectionswererehydratedfor2minindistilledwaterandthentransferredtothe0.06%potassiumpermanganatesolutionfor10min.Itisworthmentioningthattheincubationtimeinpotassiumpermanganatemayneedtobereducedwhenco-localizingthoseantigenicepitopessusceptibletochemicaloxidation.Theslideswerethenrinsedfor2minindistilledwater,transferredtotheFluoro-Jade®Cworkingsolutionfor10minandthenrinsed,airdehydrated,xyleneclearedandcoverslippedwithDPX,aspreviouslydescribed.ThebluenuclearlabelconferredbyDAPIisvisualizedviaultravioletlightexcitation,whiletheredTRITClabeledantibodyisvisualizedbygreenlightexcitation.
      BIOLOGicalInformation
      PuritySilicaTLC(acetanitrile/water,6/4)revealedthepresenceoftwofluorescentspots,presumablycorrespondingtothedisulphatehomologues.Thepresenceofprecursorsorfreefluoresceinwasnotdetected.
      PhysicochemicalInformation
      Dimensions
      MaterialsInformation
      MaterialsInformation
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