企业QQ
官方微信
FabfragmentantibodiesaregeneratedbypapaindigestionofwholeIgGantibodiestoremovetheentireFcportion,includingthehingeregion.Theseantibodiesaremonovalent,containingonlyasingleantigenbindingsite.ThemolecularweightofFabfragmentsisabout50kDa.
Storefreeze-driedpowderat2-8°C.Whenreadytouse,rehydratewithindicatedvolumeofd.waterandcentrifugeifnotclear.Productisstableforabout6weeksat2-8°Casanundilutedliquid.Prepareworkingdilutionfresheachday.Forextendedstorageafterrehydration,addanequalvolumeofglycerol(ACSgradeorbetter)forafinalconcentrationof50%,andstoreat-20°Casaliquid.Note:aftertheadditionofglycerol,theconcentrationofproteinandbuffersaltsisone-halfoftheoriginal.Alternatively,aliquotandfreezetheproductat-70°Corbelowintheabsenceofglycerol.Avoidrepeatedfreezingandthawing.
Expirationdate:oneyearfromdateofrehydration.However,theexpirationdatemaybeextendediftheproductisstoredaccordingtotherecommendationandthetestresultsareacceptableforitsintendeduse.
1:100-1:800forsequentiallabelingapplications.
Tocomplexwithprimaryantibodyinsolution,use1:1weightratioofFab:primaryantibody(15:1molarratio).Vortexandincubatefor30minutesatroomtemperaturepriortouse.Titratecomplextooptimaldilutionforassay.
Dilutionfactorsarepresentedintheformofarangebecausetheoptimaldilutionisafunctionofmanyfactors,suchasantigendensity,permeability,etc.Theactualdilutionusedmustbedeterminedempirically.
Cy3isbrighter,morephotostable,andgiveslessbackgroundthanotherorange-redfluorescingdyeconjugates.Cy3conjugatescanbeexcitedmaximallyat550nm,withpeakemissionat570nm.Forfluorescencemicroscopy,Cy3canbevisualizedwithtraditionaltetramethylrhodamine(TRITC)filtersets,sincetheexcitationandemissionspectraarenearlyidenticaltothoseofTRITC.WerecommendCy3asabrighteralternativetoTRITC.Cy3canbeexcitedtoabout50%ofmaximumwithanargonlaser(514nmor528nmlines),ortoabout75%ofmaximumwithahelium/neonlaser(543nmline)ormercurylamp(546nmline).Cy3hasbeenusedwithfluoresceinfordoublelabeling;however,theuseofanarrowband-passemissionfilterforfluoresceinisrecommendedtominimizeCy3fluorescenceintheFITCfilterset.Cy3canalsobepairedwithAlexaFluor®647formultiplelabelingwhenusingaconfocalmicroscope.However,abetterchoiceformultiplelabelingisRhodamineRed-Xbecauseitsfluorescenceismidwaybetweenagreenfluorescingdye(likeAlexaFluor®488)andafar-red-fluorescingdyelikeAlexaFluor®647.
