RatPrimaryLungMesenchymalStemCells

CatalogNo.R2303

SuggestedMedium: M5566–MesenchymalStemCellMedium/wKit-500ml

ProductDescription

RatLungPrimaryMesenchymalStemCellsarederivedfromthelungtissuesofday-1Sprague-DawleyRats.RatLungPrimaryMesenchymalStemCellsaregrowninT25tissuecultureflaskspre-coatedwithgelatin-basedsolutionfor0.5hourandincubatedinCellBiologics’CellCultureMediumfor7-15days.Culturesarethenexpanded.Priortoshipping,cellsaredetachedfromflasksandimmediatelycryo-preservedinvials.Eachvialcontainsatleast1x10⁶cellspermlandisdeliveredfrozen.RatLungPrimaryMesenchymalStemCellsarenegativeforbacteria,yeast,fungi,andmycoplasma.Cellscanbeexpanded3-6passagesbytheratioof1:2underthecellcultureconditionsspecifiedbyCellBiologics.Repeatedfreezingandthawingofcellsisnotrecommended.MouseLungMesenchymalStemCellsaretestedforexpressionofMarkersusingantibodies,CD29andCD44positivebyflowcytometry.

LaboratoryApplications

RatLungPrimaryMesenchymalStemCellscanbeusedinassaysofstandardbiochemicalproceduresperformedwithcellculturesincludeRT-PCR,Westernblotting,immunoprecipitation,orimmunofluorescentflowcytometry. 

StorageofCellBiologicsProducts

CellBiologicsshipsfrozencellsondryice.Onreceipt,immediatelytransferfrozencellstoliquidnitrogen(-180°C)untilreadyforexperimentaluse.Live-cellshipmentisalsoavailableonrequest.

Nevercanprimarycellsbekeptat-20°C.

AuthorizedUsesofCellBiologicsProducts

RatLungPrimaryMesenchymalStemCellsfromCellBiologicsaredistributedforresearchpurposesonly.Ourproductsarenotauthorizedforhumanuse,forinvitrodiagnosticprocedures,orfortherapeuticprocedures.TransferorresaleofanyCellBiologics’cellsorproductsfromthepurchasertoothermarkets,organizationsorindividualsisprohibitedbyCellBiologicswithoutthecompany’swrittenconsent. CellBiologics’TermsandConditionsmustbeacceptedbeforesubmittinganorder.

Disclaimer

AlthoughRatLungPrimaryMesenchymalStemCellsareisolatedfromlaboratorymicetestingpathogen-free,investigatorsshouldhandlethecellsthattheyreceivefromCellBiologicswithcaution,sincenotestprocedurecancompletelyguaranteetheabsenceofinfectiousagents.

WarrantyandLiability

CELLBIOLOGICS’guaranteeappliesonlytoyourpurchaseofCELLBIOLOGICS'cellswithCELLBIOLOGICS’MediaandCoatingSolutionforappropriatecellcultureandcelltestingfollowingCELLBIOLOGICS’onlineprotocolswithin35daysfromthedateofproductdelivery.

GeneralProtocolforRecoveringorFreezingPrimaryCells

Allcellcultureproceduresmustbeconductedinabio-safetycabinet.

Anyandallmedia,supplements,andreagentsmustbesterilizedbyfiltrationthrougha0.2µmfilter.

Useaseptictechniquetopreventmicrobialcontamination.

Cryo-preservedcellsmustbestoredinliquidnitrogenorseededimmediatelyuponarrival.

Medium:

ReviewtheinformationprovidedontheCellBiologics’websiteaboutappropriateculturemedia(e.g.serumandothersupplements).Usepre-warmed(37°C)cellculturemedia(30-50ML)torecovercryo-preservedcellsandwhenchangingmediaorsplittingcells.

Coatingofflasksordishes:

CoatsterileculturedishesorflaskswithGelatin-BasedCoatingSolution(CellBiologics,CatalogNo.6950)for10-30minandthenaspiratetheexcesssolutionbeforeseedingcells.

Cellrecoveryfromcryovial:

•       Quicklythawcellsincryo-vialbyincubatingthemina37°Cwaterbathfor<1minuntilthereisjustasmallbitoficeleftinthevial.

•       Promptlyremovethevialandwipeitdownwith70%ethanol.

•       Transfercellsfromthevialtoasterilecentrifugetube.Add8-10mlofpre-warmedCellBiologicsCellCultureMedium.

•       Flushthevialwithanadditional0.5-1mlofmediumtoensurecompletetransferofcellstothecentrifugetube.

•       Centrifugecellsat200gfor5minutes.

•       Aspiratethesupernatantandresuspendthecellpelletin6mlofCellBiologics’CellCultureGrowthMedium.

•       AddresuspendedcellsintoaT75flaskpre-coatedwithGelatin-BasedCoatingSolution(CellBiologics,CatalogNo.6950).

•       PlacetheT75flaskinahumidified,5%-CO₂incubatorat37°C.

•       Changeculturemediathefollowingdaytoremovenon-adherentcellsandreplenishnutrients.

•       Changecellculturemediumeverydaywhencellsare>70%confluent.

•       Cellsshouldbecheckeddailyunderamicroscopetoverifyappropriatecellmorphology.

Expansionofculturedprimarycells:

•       Removeanddiscardthecellculturemediafromtheflask.

•       Flushtheadherentlayer2timesusinga5mlsterilepipettewithsterilePBS(1X)withoutcalciumandmagnesiumtodislodgelooselyattachedcellsandremovefraction.

•       Removeanddiscardthewashsolutionfromtheflask.

•       Incubatecellswithwarm(37°C)0.05%Trypsin-EDTAsolution(CellBiologics,CatalogNo.6915)for2-5minutes.Use3.0mlofTrypsin-EDTAsolutionwhencollectingcellsfromT75flasks,and2mlwhenusingT25flasks.Assoonascellshavedetached(theflaskmayrequireafewfirmgentletaps),add8-10mlofCellBiologics’CellCultureMediumsupplementedwith5-10%FBStoaT25orT75flask(theFBSwillneutralizethetrypsin).

•       PlatecellsinfreshflasksorplatesprecoatedwithGelatin-BasedCoatingSolutioninahumidified,5%-CO₂incubatorat37°C.

•       Changeculturemediathefollowingdaytoremovenon-adherentcellsandreplenishnutrients.

•       Cellsshouldbecheckeddailyunderamicroscopytoverifyappropriatecellmorphology.

•       Changeculturemediumevery24-48hours.Pleasenotethatthemediumshouldbechangedeverydaywhencellsare>70%confluenttoremovenon-adherentcellsandreplenishnutrients.Pre-washcellswith1XPBS1-2timeswheneverreplacingthemedium.

Werecommendsplittingprimarycellsatthefollowratio:

•       Therecommendedsplitratioforprimarymurinecellsis1:2or1:3.

ProcedureforFreezingCells

Materials:

•     1XPhosphateBufferedSaline(PBS-1X)

•     0.05%Trypsin-EDTA(1X)solution(CellBiologics,CatalogNo.6915)

•     TissueCultureMedia

•     ColdFreezingMedia(10%DMSO,20%FBSand70%culturemedium,CellBiologics,CatalogNo.6916).

•     LabeledCryovials

•     Confluentcells

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