Bridge-It®S-AdenosylMethionine(SAM)FluorescenceAssayKit,384-wellformat

Application:

• QuantificationofSAMlevelinBIOLOGicalfluids,cellcultureandfermentationmedium,andextractsoftissuesandcells

• Epigeneticsandmethylationstudies

• NumerousneurologicalandpsychiatricdisordersincludingAlzheimer’sdisease,depression,HIV-relatedneurologicaldysfunction/dementia,multiplesclerosis,Parkinson’sdisease,spinalcorddegeneration,epilepsy,fibromyalgia,migraineheadaches,andalsochronicliverdysfunction,arteriosclerosisandcancer

Features:

• Easy: MixtestsampleorstandardwiththeAssaysolutionandincubateat~25°C

• Fast: Readfluorescentsignalafter30minutesofincubation

• Sensitive: AssaymeasuresSAMatalowerlimitofdetection of0.5µM (i.e.50pmol/wellina96-wellor20pmolina384-wellblackmicroplate)

• FlexIBLe: Assayisadaptabletobothlow-andhigh-throughputscreeningformats

BackgroundinformationonS-AdenosylMethionineandtheBridge-ItFluorescenceAssay

S-AdenosylMethionine S-adenosylmethionine(alsoreferredtoasSAM,SAMeorAdoMet)playsacrucialroleinthebiologicalprocessofmethylationinalltypesoforganisms.Inthemethylationcycle,SAMservesasthedonorofthemethylgroupusedinthecovalentmodificationofDNAandproteins.VariABIlityinSAMlevelshavebeenlinkedtotheprocessesofaging,numerousneurologicalandpsychiatricdisordersincludingAlzheimer’sdisease,depression,HIV-relatedneurologicaldysfunction/dementia,multiplesclerosis,Parkinson’sdisease,spinalcorddegeneration,epilepsy,fibromyalgia,migraineheadaches,andalsochronicliverdysfunction,arteriosclerosisandcancer.Currently,SAMisquantifiedusingthehighpressureliquidchromatography(HPLC)method.HPLCistimeconsuming,costlyand,duetothelargeamountoforganicsolventrequired,notenvironmentallyfriendly.

Bridge-It^® S-AdenosylMethionineFluorescence(SAM)Assay^1,2MediomicsBridge-It^® S-adenosylmethionine(SAM)fluorescenceassaymethodisbasedonacombinationofwell-establishedfluorescencemeasurementtechniquesandourpatentednewassayplatformdesignthatutilizesDNA-bindingproteinsasbiosensorsfortheirrespectivesmallmoleculeco-regulators(ligands).TheaffinityoftheDNAsequence-specificMetJproteinforitsuniqueDNA-bindingsiteisgreatlyincreasedinthepresenceofitsligand,S-adenosylmethionine.Forthisassay,theMetJconsensussequencewassplitintotwoDNA“half-sites”.OnehalffragmentwaslabeledwithfluoresceinandtheotherhalffragmentwaslabeledwithOyster^® 645fluorophore^3,4.TherelativeamountofSAMpresentinatestsamplewillinfluencetheamountofDNA-MetJproteincomplexformationintheassay.Whenthiscomplexforms,itbringsthefluorescencelabeled-DNAhalf-sitesintocloseproximityandcausesameasurablechange(increase)influorescencesignalemissionthatcanbereADIlymeasuredusingamicroplatereader(wavelengthsettings:absorption485nm;emission665nm).SAMconcentrationsintestsamplesarethendeterminedusingaSAMstandardcurve.TheBridge-It^® SAMfluorescenceassaymethodexhibitshighlydesirableperformancecharacteristicsincludingahigh(>6:1)signaltobackground(S/B)ratio,abroaddetectionrange(~0.4µM–100µM),and,adetectionsensitivityof0.4µM.Thisdetectionlevel(0.4µM)isusefulforquantifyingSAMinmosttestsamplesofinterestincludingbiologicalfluids,cellcultureandfermentationmedium,andextractsoftissuesandcells.

IncontrasttotheHPLCprocedure,theBridge-It^® SAMfluorescenceassaymethodutilizesthe384and96-wellmicroplateformat.Thus,itisideallysuitedfortherapid,simultaneousmeasurementofSAMlevelsinlargenumbersoftestsamples.Also,thismethodisreadilyadaptabletothehigh-throughputscreeningplatformscurrentlybeingusedindrugdiscoveryresearch.

Pricing: The100-wellkitisaone-timetrialpurchase.Forbulkpricingoptions,please contactus.Foranyotherquestionsorcomments,pleasedonothesitateto contactus.Wearealwayshappytohelp!

Forinformation ontheassayprinciplepleaseseethe Bridge-ItAssayTechnologyplatform pageordownloadthe protocol. Thisproductisprotectedbypatentsandpendingpatents.

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SelectedCustomerPublicationsusing Mediomics’Bridge-ItSAMFluorescenceAssayKit

1. IncreasedSerineandOne-CarbonPathwayMetabolismbyPKCλ/ιDeficiencyPromotesNeuroendocrineProstateCancer. Reina-CamposM,LinaresJF,DuranA,CordesT,L’HermitteA,BadurMG,BhangooMS,ThorsonPK,RichardsA,RooslidT,Garcia-OlmoDC,Nam-ChaSY,Salinas-SanchezAS,EngK,BeltranH,ScottDA,MetalloCM,MoscatJ,Diaz-MecoMT. CancerCell.2019Mar18;35(3):385-400.e9.doi:10.1016/j.ccell.2019.01.018.Epub2019Feb28.

2. Mitochondrialstresstriggersapro-survivalresponsethroughepigeneticmodificationsofnuclearDNA. MayorgaL,SalassaBN,MarzeseDM,LoosMA,EiroaHD,LubienieckiF,GarcíaSamartinoC,RomanoPS,RoquéM. CellMolLifeSci.2019Apr;76(7):1397-1417.doi:10.1007/s00018-019-03008-5.Epub2019Jan23.

3. PLZFtargetsdevelopmentalenhancersforactivationduringosteogenicdifferentiationofhumanmesenchymalstemcells. AgrawalSinghS,LerdrupM,GomesAR,vandeWerkenHJ,VilstrupJohansenJ,AnderssonR,SandelinA,HelinK,HansenK. Elife.2019Jan23;8.pii:e40364.doi:10.7554/eLife.40364.

4. Genescontrollingtheactivationofnaturalkillerlymphocytesareepigeneticallyremodeledinintestinalcellsfromgerm-freemice. PoupeauA,GardeC,SulekK,CitirikkayaK,TreebakJT,ArumugamM,SimarD,OlofssonLE,BäckhedF,BarrèsR. FASEBJ.2019Feb;33(2):2719-2731.doi:10.1096/fj.201800787R.Epub2018Oct10.

5. MPTACDeterminesAPPFragmentationviaSensingSulfurAminoAcidCatabolism. SuganumaT,SwansonSK,GogolM,GarrettTJ,Conkright-FinchamJ,FlorensL,WashburnMP,WorkmanJL. CellRep.2018Aug7;24(6):1585-1596.doi:10.1016/j.celrep.2018.07.013.

6. C. elegansMRP-5ExportsVitaminB12fromMothertoOffspringtoSupportEmbryonicDevelopment. NaH,PonomarovaO,GieseGE,WalhoutAJM. CellRep.2018Mar20;22(12):3126-3133.doi:10.1016/j.celrep.2018.02.100.

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1. Bridge-It^® isaregisteredtrademarkofMediomics,LLC,St.Louis,Missouri,U.S.A.Mediomicshasaworldwide,exclusivelicenseforthisassayplatformfromSaintLouisUniversity,St.Louis,Missouri.

2. TheBridge-It^® S-adenosylmethionine(SAM)fluorescenceassaykitisprovidedforlaboratoryR&Duseonly.ThisproductisnotapprovedbytheU.S.Governmentorbythegovernmentofanyothercountryoftheworldforuseinclinicaldiagnosisofdiseaseortreatmentofdiseaseinhumansoranimals.

3. Oyster^® isaregisteredtrademarkofDenovoBiolabels,GmbH,Munster,Germany.

4. Flownamics^® AnalyticalInstruments,Inc.,Madison,Wisconsin,istheauthorizedU.S.distributorofOyster®dyes.

Catalog#: 1-1-1004A(100measurements), 1-1-1004B(384measurements)

Protocol:

Bridge-It®S-AdenosylMethionine(SAM)FluorescenceAssayKit,384wellformatProtocol814.95KB

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SDS: 

S-AdenosylMethionine(SAM)FluorescenceAssayKitSDS199.12KB

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SupportingProtocol: 

ExtractionofSAMfromHEK293cells,384wellformat495.39KB

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MeasurementofSAMinBacterialCells,384wellformat197.82KB

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