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Thefetuinglycoproteinstandardwasdevelopedforuseduringglycanreleaseandlabeling(particularlyforDMBsialicacidanalysis).Fetuinexistsinavarietyofglycoformscontainingbi-,tri-,andtetra-antennaryoligosaccharideswithvariablesialylation,aswellasO-linkedglycans.
AmountofGlycoproteinSupplied
GCP-FET-05500µg
GCP-FET-250U250µg
GCP-FET-50Ux450µgpervial–4vials
SourceFetalcalfserum
FormDry.Lyophilisedpowder.
MolecularWeight36kDa(proteinweightonly)
AmountofProtein
GCP-FET-05300µg
GCP-FET-250U150µg
GCP-FET-50Ux432µgpervial–4vials
IncomparisontoBSAstandard,determinedbyBCAassay.Valueroundedtonearestμg.
Analysis
FetuinglycanswerereleasedfromtheFetuinGlycoproteinStandard(Cat#GCP-FET-05)usingPNGase-F.
Followingreleasetheglycanswerelabeledusing2-Aminobenzamide(2-AB)usingtheLudgerTag2-ABGlycanLabelingKit(Cat#LT-KAB-A2).
Figure1showsaLudgerSepN2HPLCprofileofbovinefetuinN-glycans.TothoroughlyinvestigatetheN-glycanswefirstseparatethembasedonchargeonaLudgerSepC3column(Figure2)andthenruneachfractiononaLudgerSepN2column. StorageRefrigerate(-20°C)bothbeforeandafterdissolving.Thisproductisstableforatleast5yearsassupplied. ShippingTheproductisshippedatambienttemperature. HandlingOncedissolvedavoidrepeatedthawingandrefreezing,storageover3hoursatroomtemperatureorabove,exposuretolightandlongtermexposuretoacidasthesewillcauseglycandesialylation. SafetyThisproductisnon-hazardousandhasbeenpurifiedfromnaturalsourcescertifiedtobefreeofallhazardousmaterialincludingpathogenicBIOLOGicalagents.
Figure1:WatersBEHcolumnHPLCprofileof2-ABlabeledFetuinN-glycans,releasedbyPNGaseFfromGCP-FET-05batchB1B2-04.
Figure2:LudgerSep-C3profileandsubsequentLudgerSepN2analysisofbovinefetuinPNGaseFreleasedN-glycansfromasimilarbatchofGCP-FET.
2-ABlabeledglycanswereseparatedontheLudgerSepC3columnandthesefractionswerethenseparatedontheLudgerSepN2column.ThisfiguredemonstratesthecomplexityofN-glycanspresentinthesample.AcombinationofLudgerSepC3/N2andexoglycosidasedigestionisrequiredtoidentifytheglycansandtheirrelativeabundance.N-neutralglycans,1S–monosialylatedglycans,2S–disialylatedglycans,3S–trisialylatedglycans&4S–tetrasialylatedglycans.Formoredetailedinformationonglycanidentification,pleaseseetheproductdocumentation.
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