Determinationoftheantibodytiter
Todeterminethetiteroftheantibody,anELISAwasperformedusingaserialdilutionofantibodydirectedagainstDNMT3B(Cat.No.25219).Theplateswerecoatedwiththepeptidesusedforimmunizationoftherabbit.Byplottingtheabsorbanceagainsttheantibodydilution(Figure1),thetiteroftheantibodywasestimatedtobe1:220,000.

WesternblotanalysisusingtheantibodydirectedagainstDNMT3B
Wholecellextracts(25μg)fromHeLacellswereanalysedbyWesternblotusingtheantibodyagainstDNMT3B(Cat.No.25219)diluted1:1,000inTBS-Tweencontaining5%skimmedmilk.Thepositionoftheproteinofinterestisindicatedontheright;theMarker(inkDa)isshownontheleft.

ImmunofluorescenceusingtheantibodydirectedagainstDNMT3B
HumanHeLacellswerestainedwiththeantibodyagainstDNMT3B(Cat.No.25219)andwithDAPI.Cellswerefixedwith4%formaldehydefor10minutesandblockedwithPBS/TX-100containing5%normalgoatserumand1%BSA.ThecellswereimmunofluorescentlylabelledwiththeDNMT3Bantibody(left)diluted1:1,000inblockingsolutionfollowedbyananti-rabbitantibodyconjugatedtoAlexa488.ThemiddlepanelshowsstainingofthenucleiwithDAPI.Amergeofthetwostainingsisshownontheright.

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