Background
5-hydroxymethylcytosine(5-hmC),asasixthDNAbasewithfunctionsintranscriptionregulation,hasbeendetectedtobeabundantinhumanandmousebrainandembryonicstem(ES)cells.Inmammals,itcanbegeneratedbyoxidationof5-methylcytosine(5-mC),areactionmediatedbytheten-eleventranslocation(TET)familyof5-mChydroxylases.5-hmCwasdemonstratedtoplayanimportantanddifferentrolethan5-mCintheregulationofDNAmethylation,chromatinremodeling,andgeneexpression. Concentration
1mg/ml Description
Mousemonoclonalantibodyto5-Hydroxymethylcytosine(5-hmC),cloneHMC/4D9 Specificity
Modifiedbase5-hydroxymethylcytosine(5-hmC),abroadrangeofspecies Isotype
IgG1 Formulation
10mMphosphatebuffer,150mMNaCl,pH7.4 Storage
4°C.Forlong-termstorage,aliquotandstoreat-20°C.Avoidrepeatedfreezingandthawing.Multiplefreeze/thawcyclesmayresultindecreasedperformance.Stablefor6monthsfromthedateofshipment. Purity
ProteinApurified HandlingRecommendations
Formaximumrecoveryoftheproducts,centrifugethevialpriortoopeningthecap AlternativeNames
5-hydroxymethylcytidine,anti-5-hydroxymethylcytidine,anti-5-hydroxymethylcytosine,anti-5hmC,anti-5-hmC,anti-5-hmeC,anti-5hmeC,5hmC,5hmeC,5-hmeC Application
Immunofluorescence:1:200–1:500
Immunohistochemistry:1:200–1:500
ELISA:1:1000–1:2000
DotBlot: 1:2000
hMeDIP:0.5-1µg/106Cells | 
 Fig.1.DotblotanalysisofA-1018withDNAoligoscontainingcytosine(C),5-methylcytosine(5mC),or5-hydroxymethylcytosine(5hmC).10ngofC,5mC,and5hmCDNAoligoseachwerespottedonamembrane.Themembranewasincubatedwith0.5μg/mlofEpigenteks5hmCCloneHMC/4D9antibody(1:2000dilution),thenwithperoxidaseconjugatedgoatanti-mouseIgG(dilution1:3000).Specificsignalwasonlyobservedwith5-hmCDNAoligos.
Fig.2.DemonstrationofhighsensitivityandspecificityofEpigenteks5-hmCCloneHMC/4D9antibodyagainst5-hmCDNA.SyntheticDNAoligoscontaining5-hmC,5-mCoronlycytosine,respectivelywereaddedintotheassaywellsatdifferentconcentrationsandthenspecificsignalswasonlydetectedwith5-hmCDNA.
Fig.3. SelectiveenrichmentofhydroxymethyatedDNA.50pgofunmethylated,methylated,andhydroxymethylatedDNAoligoswereeachspikedintofragmentedhumangenomicDNA(500ng).hMeDIPwasprocessedwithEpigenteks5-hmCCloneHMC/4D9antibody.ElutedDNAwasanalyzedbyrealtimePCRwiththeprimerstodetectthepresenceofspikedDNAoligos.Fold-enrichmentrepresentstheamountofrecoveredDNAoligosandwascalculatedbasedontheCts. |
