ProductDescription
Vitronectinisa478aminoacidprotein(1-19aa=signaldomain)thatbelongstoamemberofthepexinfamily.Vitronectinisanabundantglycoproteinfoundinserumandtheextracellularmatrix.ItpromotescelladhesionandspreADIng,inhibitsthemembrane-damagingeffectoftheterminalcytolyticcomplementpathway,andbindstoseveralserpinserineproteaseinhibitors.Itisasecretedproteinandexistsineitherasinglechainformoraclipped,twochainformheldtogetherbyadisulfidebond.Vitronectinhasbeenspeculatedtobeinvolvedinhemostasisandtumormalignancy.
RecentpublicationsindicatedthatcoatedrecombinanthumanvitronectinproteinalonebenefitsiPScellgenerationwhencombinedwithE8culturemedium.Theproducthasalsobeenshowntobeanexcellentcoatingmatrixmaterialfor11R-taggedrecombinantTFintracellulardeliveryforproteinderivediPSprotocolwithextremelylowlevelsofnon-specificinteraction.RecombinanthumanVitronectingene(20-398aaFragment)wasconstructedwithcodonoptimizationandexpressedinnon-fusionproteinforminE.coliasinclusionbodies.Thefinalproductwasrefoldedusingaunique“temperatureshiftinclusionbodyrefolding”technologyandchromatographicallypurified.
Vitronectinisidealforcoatingofsurfaces.Theoptimalconcentrationforcellattachmentandculturemaydifferforvariouscelltypes.Vitronectinhasbeenusedatafinalcoatingconcentrationaslowas50ng/cm2onplasticware.Further,coatingthisrecombinantproteinat5–10µg/well(6wellplateineitherNutriStemorE8mediumcanbeusedforhumaniPScellgenerationinvitro.Itisprovidedinuser-friendlypackagingforuseandstorage.Vitronectinissterilefilteredandissuppliedasareadytousesolutionafterthawingandconcentrationadjustment.
| Parameter,Testing,andMethod | RecombinantVitronectin#5121 |
| Quantity | 0.5mg |
| Volume | 1mL |
| Concentration | 0.5mg/mL |
| Purity-SDSPAGEElectrophoresis | >90% |
| Formulation | 20mMpH8Tris-HCLbufferwithproprietaryformulationofNaCl,KCl,EDTA,Arginine,DTTandGlycerol |
| Form | Solution |
| Source | Recombinant-E.Coli |
| StorageTemperature | -20°Cor-70°Cforlongtermstorage |
| ShelfLife | Minimumof6monthsfromdateofreceipt |
| SterilizationMethod | Filtration |
| CellAttachmentAssay | Passes |
| AccessionNumber | NP_000692 |
DirectionsforUse
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CoatingProcedure:
Usetheserecommendationsasguidelinestodeterminetheoptimalcoatingconditionsforyourculturesystem.
- ThawVitronectinanddilutetodesiredconcentrationusingserum-freemediumorPBS.Thefinalsolutionshouldbesufficientlydilutesothatthevolumeaddedcoversthesurfaceevenly.
- Addappropriateamountofdilutedmaterialtoculturesurface.
- Incubateatroomtemperaturefor1-2hours.
- Aspirateremainingmaterial.
- RinseplatescarefullywithdH2O–avoidscratchingbottomsurfaceofplates.
- Platesarereadyforuse.Theymayalsobestoredat2-8°Cdamporairdriedifsterilityismaintained.
ProductApplications
CoatingrecombinanthumanVTNproteinforESoriPScellculturecanbeeasilyperformedasfollowing:
1.Add1mlPBSbuffertoasinglewellof6wellplate,andmixwellwith10ugrecombinantVTN(20ulsolution,0.5mg/mlofVTNstockingsolution).
2.Placecultureplateat4°Cforovernight.
3.PlatesarereadyforroutineESculture.(RemovecoatingPBSbufferbeforecellculture).
ProductReferences
VitronectinReferences:
Wiley,LukeA.,etal."Usingpatient-specificinducedpluripotentstemcellsandwild-typemicetodevelopageneaugmentation-basedstrategytotreatCLN3-associatedretinaldegeneration."Humangenetherapy27.10(2016):835-846.
Wiley,LukeA.,etal."Generationofxeno‐free,cGMP‐compliantpatient‐specificiPSCsfromskinbiopsy."CurrentprotocolsinstemcellBIOLOGy42.1(2017):4A-12.
Dwyer,SheilaFigel,LingqiuGao,andIrwinH.Gelman."Identificationofnovelfocaladhesionkinasesubstrates:RoleforFAKinNFκBsignaling."Internationaljournalofbiologicalsciences11.4(2015):404.
Wiley,LukeA.,etal."cGMPproductionofpatient-specificiPSCsandphotoreceptorprecursorcellstotreatretinaldegenerativeblindness."Scientificreports6(2016):30742.
Kittur,Harsha,etal."ProbingCellAdhesionProfileswithaMicroscaleAdhesiveChoiceAssay."Biophysicaljournal113.8(2017):1858-1867.
Dwyer,SheilaFigel,andIrwinH.Gelman."Cross-phosphorylationandinteractionbetweenSrc/FAKandMAPKAP5/PRAKinearlyfocaladhesionscontrolscellmotility."Journalofcancerbiology&research2.1(2014).
Worthington,KristanS.,etal."Two-photonpolymerizationforproductionofhumaniPSC-derivedretinalcellgrafts."ActaBiomaterialia55(2017):385-395.
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ProductDisclaimer
ThisproductisforR&Duseonlyandisnotintendedforhumanorotheruses.PleaseconsulttheMaterialSafetyDataSheetforinformationregardinghazardsandsafehandlingpractices.
