ChIPresultsobtainedwiththeantibodydirectedagainstH3R17me2(asym)
ChIPassayswereperformedusinghumanosteosarcoma(U2OS)cells,theantibodyagainstH3R17me2(asym)(Cat.#25275)andoptimizedPCRprimersetsforqPCR.ChIPwasperformedusingshearedchromatinfrom1.6millioncellsperChIPreaction.Atitrationoftheantibodyconsistingof2,5,10and15µlperChIPexperimentwasanalyzed.IgG(5µg/IP)wasusedasnegativeIPcontrol.Figure1showstherecovery,expressedasa%ofinput(therelativeamountofimmunoprecipitatedDNAcomparedtoinputDNAafterqPCRanalysis).
Figure1A:QPCRperformedwithprimersfortheGAPDHpromoterandforexon2ofthemyoglobingene.
Figure1B:QPCRperformedwithprimersforthepromoteroftheactiveALDOAgeneandforthecodingregionoftheinactiveMYODgene.

Determinationofthetiter
Todeterminethetiter,anELISAwasperformedusingaserialdilutionoftheantibodydirectedagainstH3R17me2(asym)(Cat.#25275).Theantigenusedwasapeptidecontainingthehistonemodificationofinterest.Byplottingtheabsorbanceagainsttheantibodydilution(Figure2),thetiteroftheantibodywasestimatedtobe1:40,000. 

CrossreactivitytestusingtheantibodydirectedagainstH3R17me2(asym)
ADotBlotanalysiswasperformedtotestthecrossreactivityoftheantibodyagainstH3R17me2(asym)(Cat.#25275)withpeptidescontainingothermodificationsofhistoneH3andH4andunmodifiedsequencesfromhistoneH3.Onehundredto0.2pmolofthepeptidecontainingtherespectivehistonemodificationwerespottedonamembrane.Theantibodywasusedatadilutionof1:20,000.Figure3showsahighspecificityoftheantibodyforthemodificationofinterest.

WesternblotanalysisusingtheantibodydirectedagainstH3R17me2(asym)
HistoneextractsofHeLacells(15µg)wereanalyzedbyWesternblotusingtheantibodyagainstH3R17me2(asym)(Cat.#25275)diluted1:250inTBS-Tweencontaining5%skimmedmilk.Thepositionoftheproteinofinterestisindicatedontheleft;theMarker(inkDa)isshownontheright. 

Storeat–80°Cforupto2years.Centrifugeafterfirstthawtomaximizeproductrecovery.Aliquottoavoidrepeatedfreeze/thawcycles.Aliquotsmaybestoredat–20°Cforatleastonemonth.