ProductDescription
Glycosil®isathiol-modifiedhyaluronicacidandaconstituentofnativeextracellularmatrix(ECM).GlycosilisusedinconjunctionwithGelin-S®orECMproteinssuchaslaminin,collagen,orfibronectinformost3-Dcellcultureandtissue-engineeringapplications.Testedforbacteriaandendotoxins.GlycosilhyaluronicacidisacomponentoftheHyStem®,HyStem-CandHyStem-HPhydrogelkitsandcanbepurchasedseparatelyinindividualvials.
Glycosil®(thiol-modifiedsodiumhyaluronate)ispackagedin5mLvialscontaining50mg.Vialsareblanketedbynitrogenandunderaslightvacuum.
StoreGlycosilintheoriginalvial,unopened,at-20°Cforuptooneyear.DonotuncaptheGlycosilvialssincetheywillcrosslinkinthepresenceofoxygen.UseasyringeandneedletoaddDGWaterandremoveproductfromthevialsthroughtheseptum.
Note:Itisrecommendedtoreconstituteeachvialinitsentirety.
DirectionsforUse
GlycosilispreparedbydissolvingthelyophilizedsolidintheDGWater(oranysterile,degassed,deionizedwater).Whenre-constituted,itwillbein1xphosphatebufferedsaline(PBS),pH~7.4.TheamountofDGWaterusedfordissolutionwilldependonthevial.
Glycosilshouldbepreparedinthefollowingmanner:
- AllowtheGlycosilvialtocometoroomtemperature.
- Underasepticconditions,usingasyringeandneedle,addeither1mLor5mLofDGWatertotheGlycosilvial,dependingonvialsize(1mLor5mL).
- Placethevialhorizontallyonarockerorshaker.Itwilltake<40minutesforthesolidstofullydissolve.Warmingto37°Cand/orgentlyvortexingwillspeedupdissolvingtime.Solutionswillbeclearandslightlyviscous.
- UsewithExtralink®orExtralink-Litetoformahydrogel.
- Typically,crosslinkersareusedina1:4volumeratiowiththepolymer(i.e.1.0mLofGlycosiliscrosslinkedwith0.25mLExtralink-Lite;or0.5mLGlycosilcombinedwith0.5mLGelin-S®iscrosslinkedwith0.25mLExtralink).Note:ThegelationtimewillbeshorterforgelsnotincorporatingGelin-S.HydrogelsmadeusingonlyGlycosilwillnotsupportcellattachment.UsewithGelin-Sforcellattachment.
Glycosilreconstitutedwiththerecommendedvolumeofdegassedwaterwillbea1%solutionofthiolatedHAin1XPBSsalts.Uponreconstitution,Glycosilwillremainliquidforseveralhoursuntilcrosslinker(suchasExtralinkorPEGSSDA)isadded,atwhichpointgelationwilloccurafter20-30minuteswithnotemperatureorpHmanipulationrequired.Evenwithoutcrosslinker,agelwilleventuallyformduetoauto-crosslinking.PleaseuseGlycosilwithinfourhoursofreconstitution.DilutingGlycosilwithphosphate-bufferedsalineorcellculturemediawillincreasegelationtimeorultimatelypreventgelationalltogether.ReconstitutingGlycosilinanysolventexceptpurewaterwillalterthesaltbalanceandmayimpactgelationkineticsorgelproperties.
ProductQ&A
Globularparticleslessthan75kDashouldbeabletofreelydiffusethroughaHyStemhydrogel.
WhenreconstitutedusingDGwater,thepHofeachHyStemcomponentwillbeapproximately7.4-7.6.
Oneyearfromthedateofreceipt,ifstoredproperly.
Anysterile,deionized,degassedwatercanbesubstitutedforreconstitution.However,inordertoensureaccurateandpredictabledissolutionandgelationtimes,ourDGWaterishighlyrecommended,asitisdegassed,blanketedinargon,andhasundergonevalidationtestingwitheachHyStemcomponent.
Gelin-Sprovidescellularattachmentsiteswhenincorporatedinthehydrogel.Gelin-Sisthiol-modified,denaturedcollagenI,derivedfromeitherbovineorporcinesources.Gelin-SisincludedinallHyStem-CandHyStem-HPkits.
Gelin-Shasbeenthiol-modifiedinthesamemannerasthehyaluronaninGlycosil(orHeprasil),sothatitcovalentlycrosslinkswiththeExtralinkintheHyStemhydrogels.
Yes.Peptidesthatcontainacysteineresiduecanbeused.Thecysteineresiduemustbepresentforthepeptidetobecovalentlybondedtothehydrogelsubstrate.
Yes.ECMproteins,suchaslaminin,collagen,fibronectin,orvitronectincanbenon-covalentlyincorporatedintothehydrogelpriortocrosslinking.
HyStemhydrogelsandspongesdifferinhydrationandhomogeneity.HyStemspongesaretypicallypolymerizedhydrogelsthataresubsequentlyfreeze-dried.Theresultingspongeisafibrous,meshnetworkwithporesandnichesthatenablecellstoinfiltrateandadhere.AtrueHyStemhydrogelisanencapsulatingliquidthatpolymerizesaroundsUSPendedcellsinculture.
No.ThecomplianceofthehydrogelsissetbytheamountofExtralinkcrosslinkeradded,theconcentrationofGlycosil(orHeprasil)andGelin-Sused,andtheratioofGlycosil(orHeprasil)toGelin-S.Oncethischemicalstructureofthehydrogelisfixed,itisnotalteredbyprolongedexposuretocellculturemedium.
HyStemspongescanbeterminallysterilizedbyE-beam.HyStemhydrogelshavenotyetbeenvalidatedforusewithE-beamsterilizationmethods.HyStemhydrogelsarenotterminallysterilizedbygammairrADIation.
Gelationtimeisaffectedbymultipleaspectsofthegel’scomposition.Onewaytochangethegelationtimeofahydrogelistovarytheamountofcrosslinkerused.GelswithaloweramountofExtralinkcrosslinkerwillhavealongergelationtimethanthosewithahigheramountofcrosslinker.Changingtheamountofcrosslinkerwillproduceslightchangesingelationtime.GelationtimecanbedramaticallychangedbyvaryingtheGlycosil(orHeprasil)andGelin-Sconcentrations.ConcentratedsolutionsofGlycosil(orHeprasil)andGelin-Swillcreateasolutionwithamuchshortergelationtime.ThiscaneasilybedonebyreconstitutingthecomponentsinasmallervolumeofDGWater.Alternatively,dilutingthesecomponentsinlargervolumesofDGWaterwilldramaticallyincreasethetotaltimetoformthehydrogel.
HyStemHydrogelsarevirtuallytransparentandshouldnotinterferewithmicroscopy.
HyStemhydrogelsmaygeneratemildinflammationaspartofthebody’snaturalhealingprocessinresponsetoinjury.HyStemhydrogelsdonottriggerimmuneresponsewhenusedinvivo.(Theseproductsarenotforhumanuse)
HyStemisdegradedinvivobymatrixmetalloproteinases(Collagenases)andhyaluronidases.
Trypsin,Dipase,collagenase,andhyaluronidasehavebeenusedtohelpdetachcellsfromthesurfaceorfromwithinHyStemhydrogels.
Ingeneral,theporesizeforHyStem-CandHyStem-HPhydrogelsis~17nm.
ProductApplications
Clickonthetitleofthedesiredprotocoltolearnmore:
2DCellGrowthonHyStemHydrogels
HyStem3DCellEncapsulationforCellDeliveryApplicationsGuide
HyStem3DCellEncapsulationinhydrogelsusing96-wellplates
HyStem3DCellEncapsulationinhydrogelsusingTCInserts
EnzymeDigestionofHyStemHydrogelsforRecoveryofEncapsulatedCells
FluorescentLabelingofHyStemHydrogels
CellRecoveryfromSurfaceofHyStemHydrogels
HyStemECMIncorporation
HyStemGelationTimeVariation
HyStemStiffnessVariationProtocolfor7.5mLkit
HyStemStiffnessVariationProtocolfor12.5mLkit
ProductReferences
ReferencesforHyStem®:
Gaetani,R.,etal.(2015)EpicardialapplicationofcardiacProgenitorcellsina3D-printedgelatin/hyaluronicacidpatchpreservescardiacfunctionaftermyocardialinfarction.Biomaterials61:339-348.PMID:17335875.Prestwich,G.D.,etal.(2007)3-Dcultureinsyntheticextracellularmatrices:newtissuemodelsfordrugtoxicologyandcancerdrugdiscovery.AdvEnzymeRegul47:196-207.PMID:17335875.Shu,X.Z.,etal.(2006)Synthesisandevaluationofinjectable,insitucrosslinkablesyntheticextracellularmatricesfortissueengineering.JBiomedMaterResA79:901-912.PMID:16941590.Shu,X.Z.,etal.(2003)Disulfide-crosslinkedhyaluronan-gelatinhydrogelfilms:acovalentmimicoftheextracellularmatrixforinvitrocellgrowth.Biomaterials24:3825-3834.PMID:12818555.
S.Cai,etal.(2005)Injectableglycosaminoglycanhydrogelsforcontrolledreleaseofhumanbasicfibroblastgrowthfactor.Biomaterials,26,6054-6067.D.B.Pike,etal.(2006)Heparin-regulatedreleaseofgrowthfactorsinvitroandangiogenicresponseinvivotoimplantedhyaluronanhydrogelscontainingVEGFandbFGF.Biomaterials,27,5242–5251.G.D.Prestwich,etal.(2007)3-DCultureinSyntheticExtracellularMatrices:NewTissueModelsforDrugToxicologyandCancerDrugDiscovery.invited,Adv.Enz.Res.,inpress(2007).X.Z.Shu,etal,(2006)SynthesisandEvaluationofInjectable,InSituCrosslinkableSyntheticExtracellularMatrices(sECMs)forTissueEngineering.J.BiomedMater.Res.A,79A(4),901-912.
Shu,X.Z.,etal.(2004)Insitucrosslinkablehyaluronanhydrogelsfortissueengineering.Biomaterials25:1339-1348.PMID:14643608.Mehra,T.D.,etal.(2006)Molecularstentingwithacrosslinkedhyaluronanderivativeinhibitscollagengelcontraction.JInvestDermatol126:2202-2209.PMID:16741511.Shu,X.Z.,etal.(2004)AttachmentandspreadingoffibroblastsonanRGDpeptide-modifiedinjectablehyaluronanhydrogel.JBiomedMaterResA68:365-375.PMID:14704979.Ghosh,K.,etal.(2007)CelladaptationtoaphysiologicallyrelevantECMmimicwithdifferentviscoelasticproperties.Biomaterials28:671-679.PMID:17049594.
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ProductDisclaimer
ThisproductisforR&Duseonlyandisnotintendedforhumanorotheruses.PleaseconsulttheMaterialSafetyDataSheetforinformationregardinghazardsandsafehandlingpractices.
