[DevelopmentBackgroundandFeatures]
S-CloneSF-O3isacompletelyserum-freemediumforuseinresearchofmousehematopoieticstemcells.TheproductwasdevelopedbasedupontheresultsofresearchbytheNishikawaLaboratory,DepartmentofMolecularGeneticsattheGraduateSchoolofMedicine,KyotoUniversity.Itisusedformolecularstudiesonproliferationandmaintenancemechanismsofmousehematopoieticstemcellsandhasbeenreportedeffectiveinthesame.
Themediumisserum-free,whicheliminateseffectsofanyunknownfactorsinserum,anditcontainsextremelylimitedproteincomponents,allowingtheactionofcytokines,etc.tobeclearlyidentified.Further,beingcomposedofknowningredients,theproductaffordsconstancyinexperimentalconditions.(SeeReferenceLiteratures:(1),(2),(3),(4),(5))
[Formulation]
1.Basalmedium(powder)
ConstitutedfromacompositeofslightlymodifiedRPMI1640,Dulbecco"sMEM,andHam"sF-12.
2.Sodiumhydrogencarbonate(powder)
3.Supplement(liquid)
HEPESbuffercontaininghormonegrowthfactors(insulin,transferrin,sodiumseleniteandethanolamine).Proteincomponentsotherthaninsulinandtransferrinhavenotbeenadded.
[DirectionsforUse]
(PreparingtheMedium)
1.Add1bottleofthebasalmedium(10.7g)toapproximately950mLofpurifiedwaterandmixwelltodissolve.
2.Tothesolutionabove,add1vial(2.2g)ofsodiumhydrogencarbonateanddissolve,andfurtherdilutetoatotal1,000mL.
3.Sterilizebyfilteringthroughalowadsorptionmembranefilterofporesize0.2-0.22um(ToyoRoshiKaishaLtd.,Cat.No.C020A047AorMilliporeCorporation,Cat.No.GVWP04700,etc.).
4.Asepticallyadd1vialofthesupplement(10mL)tothefiltered1,000mLofmedium.
5.Sealandstorerefrigerated(2-10C).
(ExamplesofUsage)
1.MethodforcultivationofBprecursorcellsonamousestromalcelllineST2layerusingSF-O3(AddIL-7:10U/mL,2-mercaptoethanol:100umol/L).
1)SeedtheST2cellsatadensityof1.5x10^5cells/welltomakethefeederlayer.
2)Seedthemousebonemarrowcellsatadensityof2x10^5cells/wellontheST2feederlayerandcontinuetoincubate,replacinghalfofthemediumevery4-5days.Wellused:FalconPrimariaCultureware6-WellPlate(3846)
2.MethodforcultivationofhematopoieticprecursorcellsinamousestromalcelllinePA6layerusingSF-O3(AddbFGForEGF:5-10ng/mL,2-mercaptoethanol:100umol/L).
1)SeedthePA6cellsatadensityof2x10^5cells/welltomakethefeederlayer.
2)Seedthemousebonemarrowcellsatadensityof2x10^5cells/wellonthePA6feederlayerandcontinuetoincubate,replacinghalfofthemediumevery4-5days.Wellused:FalconPrimariaCultureware6-WellPlate(3846)
[PrecautionsforUseandHandling]
1.Theentirebottleofbasalmediumshouldbedissolvedinasingleuse.Avoiddissolvingthepowderinsmallportion.
2.Whendissolvingthebasalmedium,injectcarbondioxidegas(high-qualitycarbondioxideordryiceissuitable)toshiftthepHofthemediumtoacidic.AnacidicpHenhancessolubilityofthemediumaswellasspeedsequilibration(pH7.2-7.4)ofthemediuminthecarbondioxidegasincubator.
3.Asthesupplementisaconcentratedsolution,itmayshowprecipitationwhenstoredrefrigerated.Itshouldbemixedwellbeforeaddingtothebasalmedium.
4.Avoidfilteringthemediumafterthesupplementhasbeenadded,asitmaycauseloweredgrowthinsometypesofcells.
5.Thepreparedmediumisconfirmedtobestablefor3monthswhenstoredat2-10C,however,itshouldbeusedassoonaspossIBLe.
6.Thisproductdoesnotcontain2-mercaptoethanol.
7.Thisproductisareagentforinvitroresearchuseonlyandshouldnotbeusedfordiagnosticpurposes,norshouldthegrowncellsbeusedfortreatment.
8.Treatmentofwastesgeneratedbythismethodshouldbedoneaccordingtoguidelinesinaccordancewiththelocallawsandregulations.
**Note)Pleasealsorefertothe"S-CloneSF-O3"MaterialSafetyDataSheet(MSDS)beforeuse.
MSDSforSF-O3Cat#1303(PDFFile)
[StorageandShelfLife]
*Storeat2-10C,avoidfreezing.
*Theshelflifeis1yearfollowingmanufacture(expirationdateisindicatedontheouterbox)
[PackageContents]
*Basalmedium:1L-equivalentx1bottle
*Sodiumhydrogencarbonate:2.2gx1vial
*Supplement:10mLx1unitvial
[ReferenceLiterature]
(1)ImmunologyLetters.40:163-169,1994
(2)SeminarsinIMMUNOLOGY,7:185-196,1995
(3)J.Exp.Med.,182:315-323,1995
(4)ClinicalImmunology,27(10):1193-1198,1995
(5)JournalofCellBIOLOGy,Volume132,Numbers1&2,January199691-99
(6)NatureBiotechnology23,1542-1550(2005)
(7)Cell138,300-313,July24,2009
(8)Blood2010116:4474-4482
(9)JCellSci122,3303-3311,September15,2009
(10)Development2013Jan1;140(1):216-25
ProductInsertSF-O3Cat#1303(PDFFile)
FORRESEARCHUSEONLY,SHOULDNOTBEUSEDFORDIAGNOSTICPURPOSE,NORSHOULDTHEGROWNCELLSBEUSEDFORTREATMENT.
Aserum-freemediumforuseinresearchofhematopoieticstemcells
Manufacturedby:SekisuiMedicalCo.,LTD
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