June22011July202011ABSTRACTWeshowthatalabelfreeelectrochemicalmethodcanbeusedtomonitorthepositionofanenrichedanalytebandduringbipolarelectrodefocusinginamicrouidicdeviceThemethodreliesonformationofa ID: 860925
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1 Label-FreeElectrochemicalMonitoringofCon
Label-FreeElectrochemicalMonitoringofConcentrationEnrichmentduringBipolarElectrodeFocusingEoinSheridan,DzmitryHlushkou,RobbynK.Anand,DerekR.Laws,UlrichTallarek,*RichardM.Crooks*DepartmentofChemistryandBiochemistry,CenterforElectrochemistry,andtheCenterforNano-andMolecularScienceandTechnology,TheUniversityofTexasatAustin,1UniversityStation,A5300,Austin,Texas78712-0165,UnitedStatesDepartmentofChemistry,Philipps-UniversitatMarburg,Hans-Meerwein-Strasse,35032Marburg,GermanySupportingInformationnpreviousreportswedemonstratedthatchargedanalytescanbeconcentratedupto500000-foldusingamethodwecallbipolarelectrodefocusing(BEF).Intheseearlierstudiestheconcentratedanalytewasdetectedusinguorescence,buthereweshowthatelectrochemicalmethodsarealsocapableofdetectingenrichedbandsofchargedspecies.Additionally,weprovidetheoreticalandexperimentalinsightsintothenatureoftheobservedelectrochemicalsignal.Thesignicantachievementisthatbothenrichmentanddetectionarenowelectrochemical,therebyeliminatingtheneedforuorescentlabelinganddetec-tion.Importantly,theanalytebeingdetecteddoesnotitselfneedtobeelectroactive.ThebasisofBEFismanipulationofthelocalelectricwithinanelectrolytesolutioninthevicinityofabipolarelectrodeInbrief,applicationofavoltage()acrossabulledmicrochanneloflength,resultsinanelectrichavingaconstantvalueof.IfaBPEispresentinthechannel(Scheme1a),andifissucientlylarge,faradaicelectrochemicalreactionswilloccuratthepolesoftheBPE(Scheme1b).ThissituationissatisedwhenthefractionofdroppedoverthelengthoftheBPE(whereisthelengthoftheBPE)exceedsthatrequiredtodriveareductionreactionatthecathodicpoleandanoxidationattheanodicpole.ThekeypointisthattheinterfacialpotentialbetweenthepolesoftheBPEandthesolutionisdeterminedby.ThepotentialoftheBPEisalsoafunctionofthereforewilladjusttothesurroundingsolutionpotential.ElectrochemicalprocessesmayresultinsubstantialchangestotheionicstrengthofthebuerneartheBPE.Forexample,intheexperimentsreportedhere,thebuerisTris.WateroxidationattheBPEcathodegeneratesOH,whichneutralizesTrisH,therebydepletingchargecarriersinsolutionandcreatinganelectricgradientneartheBPE.AdditionalinformationaboutthefaradaicreactionsoccurringattheBPEareprovidedintheSupportingInformation.Enrichmentofanalyteanionsoccursatthepositionontheelectriceldgradientwherethevelocityduetototal(nearlyuniformalongthelengthofthechannel)isexactlybalancedbyanequalandoppositeelectrophoreticvelocity(whichisafunctionoflocationalongtheelectriceldgradient),wheretotalowisduetoelectroosmosis(EOF)andpressure-driven(PDF)combined.ThisprocessisillustratedinScheme1c.uidic-basedsensorshavingmultiple,integratedfunctionsarenowcommonplace.However,insomecasesthelimitofdetectionofthesesystemsiscompromisedbysmallanalysisvolumesandlowanalyteconcentrations.Oneapproachforaddressingthisprobleminvolvesenrichmentoftheanalyteataspeciedlocation,forexample,nearthedetector.Manyapproachesforsampleenrichmenthavebeenreported,includingelectrokineticmembraneltration,11,12solid-phaseextraction(SPE),13,14isoelectricfocusing(IEF),andtemperaturegradientfocusing(TGF).18,19BEFisaformofelectriceldgradientfocusing(EFGF),anapproachformicrouidicconcentrationenrichmentthatisfrequentlyusedinchip-basedanalyticaldevices.Enrichmentfactorsofupto10havebeenreportedusingothermembersoftheEFGFfamily. June2,2011July20,2011 ABSTRACT:Weshowthatalabel-freeelectrochemicalmeth-odcanbeusedtomonitorthepositionofanenrichedanalytebanddur
2 ingbipolarelectrodefocusinginamicrouidic
ingbipolarelectrodefocusinginamicrouidicdevice.Themethodreliesonformationofadepletedbuercationregion,whichisresponsibleforconcentrationenrichmentofthechargedanalyte.However,thisdepletionregionalsoleadstoanincreaseinthelocalelectriceldinthesolutionnearabipolarelectrode(BPE),andthisinturnresultsinenhancedfaradaicreactions(oxidationandreductionofwater)attheBPE.Therefore,itispossibletodetectthepresenceoftheconcentratedanalytebandbymeasuringthecurrentpassingthroughtheBPEusedforconcentrationenrichment,ortheconcentratedbandcanbedetectedatasecondaryBPEdedicatedtothatpurpose.Bothexperimentsandsimulationsarepresentedthatfullyelucidatetheunderlyingphenomenonresponsiblefortheseobservations. HerewepresentamethodformonitoringthelocationofananalytebandenrichedusingBEFbymeasuringthecurrentpassingthroughtheBPE(BPE).Specically,enrichmentisinitiatedbyconnectingthetwopolesofasplitBPEthroughanammeterandthenapplyingasucientlyhightottoinitiateenrichmentofananionicuorescenttracer(Scheme1).ThiscongurationmakesitpossibletosimultaneouslymonitorthelocationoftheenrichedbandusingbothuorescencemicroscopyandBPE.Also,addi-tionalmicrobandelectrodespresentwithinthechannelmakeitpossibletoquantitativelymeasuretheelectriceldgradientinthevicinityoftheBPE.Thisprovidesinsightintotheunderlyingprinciplesofelectrochemicaldetection.Numericalsimulationsareinaccordwiththeexperimentalresults.EXPERIMENTALSECTIONPoly(dimethylsiloxane)(PDMS)channelswerepreparedusingaSylgard184elastomerkitobtainedfromK.R.Anderson,Inc.(MorganHill,CA).Au-coatedglassslides(100nmthick,noadhesionlayer)werepurchasedfromEMFCorp.(Ithaca,NY).Thefluorescenttracerwas4,4-difluoro-1,3,5,7,8-pentamethyl-4-bora-3a,4a-diaza--indacene-2,6-disulfo-nicacid,disodiumsalt(BODIPY,InvitrogenCorp.,Carlsbad,CA).PluronicF-108wasobtainedfromBASF(FlorhamPark,NJ).Trisbuffer(pH8.1)waspreparedbydilutionofa1.0MTrisHClsolutionpurchasedfromFisher(FairLawn,NJ).Photoresist(AZ4620)anddeveloper(AZ421K)werepur-chasedfromAZElectronicMaterials(Somerville,NJ).Allchemicalswereusedasreceived.Deionizedwaterhavingaresistivitygreaterthan18Mcmwasusedforallexperiments(Milli-Qgradientsystem,Millipore,Bedford,MA).MicrofluidicDeviceFabrication.Devicefabricationhasbeendescribedpreviously.Briefly,thedrivingelectrodesandBPEsweredepositedontotheglassbaseofthemicrofluidicdeviceusingstandardphotolithographictechniques.Thedrivingelec-trodesconsistedofmicrofabricatedAudiskslocatedatthebottomsofthetworeservoirs.Theywereconnectedtoapowersupplyviaexternalcontacts.Anarrayof15goldmicrobandelectrodesconsistingof30mwidelinesand50mwidespaceswasusedtoformtheBPEsandmeasuretheelectricfieldgradient.ThemicrobandshadexternalcontactssothatanytwocouldbeconnectedtoformaBPE.MicrochannelswerefabricatedinPDMSusingapreviouslydescribedreplicamoldingprocedure.y,amicrochannel(6mmlong,100mwide,and21mhigh)spanningtwo5.0mmdiameterreservoirswasfabricatedfromPDMS.ThePDMSwasrinsedwithethanolanddriedunderN,andnextboththePDMSandtheglassslidesupportingtheAuelectrodeswereexposedtoanOplasma(60W,modelPDC-32G,HarrickScientic,Ossining,NY)for30sonthemediumpowersetting.AfterjoiningthePDMSreplicaandtheglassslide,theentireassemblywasplacedinanovenat70Cfor7mintopromoteirreversiblebonding.cationofthechannelwallswithPluronicwascarriedoutusingapreviouslyreportedmethod.cally,themicrouidicdeviceswereloadedwitha3.0M,buPluronicsolutionandallowedtoequilibratefor20hat23C.Thechannelswerethenthoroughlyri
3 nsedwithpurebusolutionandusedimmediately
nsedwithpurebusolutionandusedimmediatelywithoutfurthertreatment.Mod-cationofthechannelwallswithPluronicloweredEOFmobility)from4.5to3.6/(Vs),suchthatwhen25VwasappliedtheEOFvelocity()was15InstrumentationandDataAcquisition.FluorescenceimageswereobtainedusingaNikonAZ100(NikonCo.,Tokyo,Japan)microscopeequippedwithamercurylamp(Nikon)andaCCDcamera(Cascade,PhotometricsLtd.,Tucson,AZ).Micro-graphswereprocessedusingV++PrecisionDigitalImagingsoftware(DigitalOptics,Auckland,NewZealand).Imageswerecapturedusing11binningwith512290pixelsanda100msexposuretime.Fluorescenceintensitiesforcalibrationcurveswereobtainedusingafunctionthataveragesintensitiesofallpixelsalongoneaxisoftheregionofinterest.Fluorescenceintensitiesofconcentratedanalyteswereobtainedusingamax-imumfunction,whichprovidesthemaximumintensityvaluesalongoneaxisoftheregionofinterest.ConcentrationEnrichmentExperiments.Priortoeachen-richmentexperiment,themicrofluidicchannelwasrinsedbyintroducing80.0Lof5.0mMTrisbuffer(pH8.1)intotheanodicreservoirand15.0Lintothecathodicreservoir.Thebuffersolutionwasallowedtoflowthroughthemicrochannelfor20mininresponsetothesolutionheightdifferential.Next,therinsingsolutionineachofthereservoirswasreplacedwith80.0Lof1.0 Scheme1 in5.0mMTris.AdditionalmicroliterincrementsofthesamesolutionwereaddedtoeachreservoirinindividualexperimentsasindicatedintheResultsandDiscussionsection.Theconcentrationenrichmentexperimentsthemselveswerecarriedoutasfollows.First,twomicrobandelectrodeshavingthedesiredseparation(typically500m)wereconnectedviaaconductivewireorammeter.Second,adrivingvoltage(25.0V)wasappliedacrossthemicrochannelusingahigh-voltagepowersupply(LLS9120,TDK-LambdaAmericas,Inc.,SanDiego,CA)connectedtothemicrofabricatedgolddrivingelectrodesspanningthebottomsofthereservoirs.Measurementsuorescencewereobtainedasafunctionoftime.ThePDFrateinthemicrochannelwasestimatedaftereachexperi-mentbyswitchingoandobservingthemovementoftheenrichedbandundertheinuenceofPDFonly.CurrentMeasurements.ThevalueofBPEwasmeasuredbyconnectingthepairofmicrobandsdefiningtheBPEviaanammeter(model6517Belectrometer,KeithleyInstruments,Inc.,Cleveland,OH).DatawereprocessedusingLabViewsoft-ware(NationalInstruments,Austin,TX).Thetotalcurrentflowingthroughthechannel(tot)wasmeasuredinparallelwithBPEbymonitoringthevoltagedropacrossa523kresistorplacedinserieswiththemicrochannel.Thevoltagewasmeasuredusingahand-held,digitalmultimeterequippedwithPC-Linksoftware(VA18B,SinometerInstruments,ShenZhen,China).ElectricFieldProfileMeasurements.Theaxialelectricfieldprofilewithinthechannelwasmonitoredusingascanningdigitalmultimeter(SDMM,model2700,KeithleyInstruments,Inc.,Cleveland,OH)equippedwithamultiplexermodule(model7701,Keithley)connectedtoallthemicrobandelectrodesexceptthosedefiningtheBPE.TheSDMMwascontrolledwithMicro-softExcelviathesoftwareprovidedbytheSDMMmanufacturer(ExceLinx,Keithley).TheSDMMwasinterfacedtothemicro-bandelectrodesthroughabreakoutboard(screwterminals).TheSDMMreadsthedifferencebetweenpairsofmicrobandsinsequence.Theacquisitiontimeforeachvoltagemeasurementwas0.1s,andthevoltagebetweenpairsofmicrobandswasreadevery2.0s.Electricfieldmonitoringexperimentsproceededasfollows.First,thetwohalvesoftheBPEwereconnectedviaaconductivewireorammeter.Second,theSDMMwasplacedintoscanmode.Third,adrivingvoltage(tot=25.0V)wasappliedacrossthemicrochannelviathedrivingelectrodes.Thecaptureddatawerestoredandplottedasvolta
4 gedifferencesbetweeneachmicrobandpairver
gedifferencesbetweeneachmicrobandpairversustimeinrealtimeusingExcel.ComputerSimulations.Numericalsimulationswereusedtomodelthelocalelectricfieldandspeciesconcentrationdistribu-tionsinthemicrochannel.ThebasicequationsgoverningthephysicalphenomenainthesystemarethecoupledNernstPlanck(eq1),Poisson(eq2),andNavierStokes(eq3)equations. cit¼3Diciþ ziFRT3ðDiivrið1Þ2 0rizicið2ÞF Here,istheconcentrationofspecies(mol/mitsdiusioncoecientandvalency,respectively;isthelocalelectricpotential;,andrepresenttheFaradayconstant,molargasconstant,andabsolutetemperature,respectively;owvelocity,isthereactiontermforspeciesthevacuumpermittivityanddielectricconstant;arethemassdensityanddynamicviscosityoftheliquid;ishydrostaticpressure.TheNernstPlanckequationdescribeslocalmassandchargetransportduetoadvection,diusion,andelectromigra-tion;thePoissonequationestablishesarelationshipbetweenthelocalelectricalpotentialandthespeciesconcentrationdistribu-tion;theNavierStokesequationrelatestheowvelocityeldtothepressureandLorenzforce.ThemicrochannelandreservoirsareassumedtobelledwithMBODIPYin5.0mMTrisHClbuer(pH8.1).Thus,thespeciesofinterestincludeneutralTris,itsprotonatedform(TrisH),Cl,OH,andBODIPY.Thereactiontermsineq1takeintoaccountthefactthatthelocalconcentra-tionofTris,TrisH,OH,andHcanchangeduetothebulkerreactionsand/orfaradaicreactions.Theaboveformaldescriptionoftheprocessesinthesystemwasimplementedasaniterativenumericalschemebasedondiscretespatiotemporalschemesoptimizedforparallelcomputa-tions.Inparticular,forthesolutionoftheNavierStokes,Poisson,andNernstPlanckequations,thelattice-BoltzmannapproachandthenumericalapproachesdescribedbyWarrenandbyCapuanietal.wereimplemented.Inallnumericalschemes,atimestepof10sandaspacestepof10mwereused.ThekineticsoftheassumedfaradaicandbuerreactionsusedinthesimulationsalongwithotherdetailsofthenumericalimplementationareprovidedintheSupportingInformation.Asinglesimulationrequired14husing64processorsofanSGIAltix4700supercomputertoanalyzethetemporalbehaviorofthesystemfor100s.RESULTSANDDISCUSSIONRelationshipbetweenBPECurrentandEnrichedBandThedeviceusedforenrichmentstudiesconsistedofa6mmlongmicrochannelcontaininganarrayofgoldmicrobands(Figure1a).Eachmicrobandwasconnectedtoanindividualcontactpadexternaltothedevice.BPEsofarbitrarylengthcouldthenbeformedbyplacingajumperwireorammeteracrosstheexternalcontactpads.Inthisexample,a500mlongBPEwascreatedbyconnectingtwomicrobandsthroughanammetersothatcouldbemeasured(Figure1b)throughouttheexperiment.Theinuenceoftheenrichedbandonwasinvestigatedasfollows.First,themicrochannelwaslledwith5.0mMTrisercontaining1.0MBODIPY,andinitially(=0s,Figure1b)anexcess(22.0L)ofthissolutionwasaddedtotheanodicreservoir.Second,at=50savalueof=25.0Vwasappliedtothedrivingelectrodes.Atthispoint,thePDFrateresultingfromthepresenceofunequallevelsofsolutioninthetworeservoirsresultsinatotalowratetowardthecathodicreservoirthatistoofasttoallowastabledepletionregiontoforminthevicinityoftheBPEcathode.TheSupportingInformationprovidesmoreinformationaboutthissituation,butthemainpointisthatintheabsenceofadepletionregionenrichmentdoesnotoccur.Thisinitialconditionisimportant,becauseitmakesitpossibletoobservetheimpactofinitiatingenrichmentonwithoutchanging.Figure1bshowsthatimmediatelyuponapplicationof=25.0VanincreaseofBPE60nAisobserved.Thiscurrentarisesfromoxidationandreductionof waterattheBPEanodeandcathode,respectively(seet
5 heSupportingInformation).Next,at=120s,th
heSupportingInformation).Next,at=120s,theoriginalexcessofsolutionintheanodicreservoirwaspartiallyosetbyaddingLtothecathodicreservoir.Thisresultsinadecreaseintheowratefrom245to205m/s.Coincidentwiththisdecrease,enrichmentofBODIPYcommencedandcreasedfrom55to190nA.Theunderlyingreasonforthisincreaseisthemainpointofthisstudyandwillbediscussedinthenextsubsection.FurtherchangestothePDFrate,initiatedwithadditionalincrementsofsolutiontothecathodicreservoir,resultinprogressivechangesinandmovementoftheenrichedbandawayfromtheBPE.Specically,2.0Laliquotsofsolutionwereaddedtothecathodicreservoirevery60sstartingat=220sandcontinuinguntil460s.EachaliquotcorrespondstoadecreaseinthePDFrateofm/sandresultsintheenrichedbandmovingmclosertotheanodicreservoir.Acorrespondingseriesofstepwisedecreasesinareobserved(Figure1b).Note,however,thattheprecisechangeinthetotalowrateandmovementoftheenrichedbandvariesslightlywitheachadditionofbuerduetoimperfectlyshapedreservoirsandchangesinelectroosmoticowrateinducedbythePDF.Thisisaminorpoint,butneverthelessacompletediscussionisprovidedintheSupportingInformation(FigureS1).TodemonstratethatchangesinthepositionoftheenrichedbandandBPEarereversible,theforegoingexperimentwasreversedbyadding2.0Laliquotsofsolutiontotheanodicreservoirevery60sstartingat=460s.Thisresultsintheenrichedbandmoving200mclosertotheBPEforeach2.0LadditionandstepwiseincreasesinBPE(Figure1b).ThekeytrendsillustratedinFigure1barethattheformationoftheenrichedbandresultsinasharpincreaseinBPE,andthatBPEdecreasesandincreasesastheenrichedbandmovesawayfromandtowardtheBPE,respectively.Onenalpoint:theBODIPYisitselfnotelectroactiveovertherangeofpotentialsrelevanttothisstudy,andthereforeitdoesnotcontributedirectlyFigure1cshowstherelationshipbetweenthepositionoftheenrichedBODIPYbandand,measuredoverthecourseofeightexperimentsandusingdierentmicrouidicdevices.Here,thehorizontalaxisrepresentsthedistancebetweenthecathodicedgeoftheBPEandtheedgeoftheenrichedbandclosesttotheBPE.ThesedatademonstratetheconsistentincreaseinrecordedastheenrichedBODIPYbandmovesclosertotheBPE.Accordingly,canbeusedtoestimatethelongitudinalpositionoftheenrichedband.AsdiscussedintheSupportingInformation,however,thereisnorelationshipbetweenthemagnitudeofandtheconcentrationoftheenrichedband(SupportingInformationFigureS2).ElucidationoftheSourceofBPECurrentChanges.viouslywereportedthatwaterreductionatthecathodicpoleoftheBPEresultsinneutralizationofTrisHbuffercationsaccordingtoeqs4and5.25VTheresultingformationofaregionoflowerconductivity(depletionregion)iscriticaltotheformationofasteepelectricfieldgradientandthesubsequentenrichmentprocess(Scheme1).WehavepreviouslyshownthatthepotentialdifferencebetweenpairsofmicrobandssituatedinthevicinityoftheBPE(Figure1a)canbeusedtoconstructamapofthisgradient.ThedashedlineinFigure2ashowsthatwhenthetotalflowrateistoofastforenrichmenttocommencethereis Figure1.(a)OpticalmicrographofgoldmicrobandelectrodesthatcanbeconnectedexternaltothechanneltoyieldaBPEofarbitrarylength.ThecurrentthroughtheBPE()canbemonitoredbyconnectingthetwopolesoftheBPEwithanammeter(A).(b)asafunctionoftimewhileanenrichedBODIPYbandisformedandmovedlongitudinallyinthechannelbychangingthetotalowrate.(c)RelationshipbetweenBPEandtheenrichedBODIPYbandposition.Theinitialconcentra-tionsinthemicrochannelwere5.0mMTrisHCl(pH8.1)and1.0.TheBPEhadalengthof500 essentiallynofieldgradient.However,whentheflowrateisreduced
6 (bydecreasingthePDFrate),therebyinitiati
(bydecreasingthePDFrate),therebyinitiatingenrich-ment,agradientdevelops(solidlineinFigure2a).Significantly,partoftheelevatedelectricfieldisdirectlyabovetheBPEminFigure2a).Thismeansthatishigherafterenrichmentisobserved,andthereforeagreaterdrivingforcefortheoxidationandreductionofwaterexistsatthepolesoftheBPE(Figure2b).Thisobservationexplainsthesharpincreaseinupontheinitiationofenrichment(Figure1b).Numericalsimulationswereusedtoconrmthelocationsofdepletionandtheelevatedelectriceld.SpeciFigure3ashowsthesimulatedTrisHconcentrationproleinthemicrochannel.Duringenrichment,thedepletionregionislocatedovertheBPEandjusttoitsright.TheelevatedTrisHconcentrationovertheanodicpoleoftheBPEarisesfromprotonationofTrisduetooxidationofwater(protonfor-mation).Figure3aalsoshowsthatthelocationofTrisHdepletionisafunctionofthetotalowrate(i.e.,PDF+EOF).Therefore,whenthedepletionregionmoves,thelocationoftheeldgradient(Figure3b),andhencethelocationofenrichment,alsomoves.Additionally,Figure3bshowsthatwhentheenrichedbandmovesawayfromtheBPE,thetotalelectriceldstrengthovertheBPEdecreases(lower),whichexplainsthedecreaseinrecordeduponmovingtheenrichedbandawayfromtheBPE(Figure1b).Thesimulatedelectriceldpro(Figure3b,solidlines)arequalitativelysimilartothosedeter-minedexperimentally(Figure3b,dashedlines)foragiventotalowrate.Quantitativedierencesaretheresultofnecessaryapproximationsusedforthesimulations,andthesearediscussedindetailintheSupportingInformation.Furtherevidencefortheformationandmovementofthedepletionregioncanbeobtainedbyexaminingthetotalcurrentowingthroughthemicrochannel().Figure4showstotmeasuredasafunctionoftimeduringanenrichmentexperiment.=0s,tot=25.0Vwasappliedresultingin=380nA.Atthistime,thetotalowratetowardthecathodicreservoirwastoofastforenrichmenttooccur(similartotheinitialconditionsusedinFigure1b),soBPEisquitelow(20nA).Next,at=60s,enrichmentwasinitiatedbydecreasingthePDFrate.Thisresultsinanincreasein,justaswasnotedinFigure1b.Simultaneously,totdecreasesduetothesignicantincreaseinsolutionresistance)causedbytheformationofthedepletionregionattheBPE.Next,between=100and175s,theenrichedbandwasmovedtowardtheanodicreservoirbysequentialadditionsofsolutionto Figure2.(a)MeasuredelectriceldprolesinthemicrochannelintheregionneartheBPEuponenrichmentofBODIPYbyapplicationof=25.0V.Prolesweremeasuredbefore(dashedline)andafter(solidline)enrichmentbymodulatingtheowrate.Errorbarsindicatethestandarddeviationfromthemeanforthreereplicatemeasurements.(b)Representationofthechangeinuponinitiationofenrich-ment.DepletionofTrisHinthevicinityoftheBPEresultsinahighersolutionresistanceandhenceanincreasein.Theinitialconcen-trationsinthemicrochannelwere5.0mMTrisHCl(pH8.1)and1.0MBODIPY.TheBPEhadalengthof500 Figure3.(a)SimulatedTrisHconcentrationproles.(b)Simulated(solidline)andmeasured(dashedline)electriceldprolesduringaenrichmentexperiment.TheBPEspanstherangefrom0tom.Theinitialconcentrationsinthemicrochannelwere5.0mMHCl(pH8.1)and1.0MBODIPY=25.0V. thecathodicreservoir.EachadditionofsolutionresultsinadecreaseBPE.Simultaneously,totdecreasesduetotheexpansionofthedepletionregionastheenrichedbandmovestowardtheanodicreservoir(Figure3a).ThetemporalrelationshipofthechangesinBPEinFigure4provideevidenceoftheresponsivenessofBPEtotheformationandmovementofanenrichedband.EffectofEnrichedBandPositiononaSecondaryBPE.furtherillustratetheabilityofthedepletionregiontomodulate,itsinfluenceonthecurr
7 entpassingthroughasecondaryBPEwasmonitor
entpassingthroughasecondaryBPEwasmonitored.Fortheseexperiments,thesecondaryBPEwasplacedbetweentheenrichmentBPEandtheanodicreservoirbyconnectingasecondpairofmicrobands(Figure5a).Figure5bforthesecondaryBPEwhenanenrichedband(formedattheenrichmentBPE)ismovedoverthesecondaryBPEandtotherightofitbychangingthePDFrate.FaradaicreactionswereinducedinthesecondaryBPEwhentheenrichedbandwasaboveortotherightofthesecondaryBPE(=50s),andtheywereswitchedoffagainwhentheenrichedbandwaspushedbacktotheleftofthesecondaryBPE(=175s).WhentheenrichedbandwaslocatedbetweentheenrichmentBPEandthesecondaryBPE(Figure5d,solidline),theelectriceldtotherightoftheenrichedband(includingthesecondaryBPEregion)wasrelativelylowinmagnitude.ThesecondaryBPEexperiencedanelectriceldstrengthof4.0kV/mcorrespond-ingtoa1.44VforthesecondaryBPE(360minlength).ThisisinsucienttoinducefaradaicreactionsatthesecondaryBPE(seetheSupportingInformation).However,asthepositionoftheenrichedbandispushedfurthertotherightsuchthattheenrichedbandislocatedoverthesecondaryBPE Figure4.(solidline)and(dashedline)asafunctionoftimeforanenrichedBODIPYbandbeingformedandmovedtowardtheanodicreservoirbymodulationofthetotalowrate.Theinitialconcentrationsinthemicrochannelwere5.0mMTrisHCl(pH8.1)and1.0MBODIPY.TheBPEhadalengthof500=25.0V. Figure5.(a)OpticalmicrographshowingadevicecontaininganenrichmentBPEandasecondarydetectionBPE.(b)BPEasafunctionoftimeshowingtheincreaseincurrentthroughthesecondaryBPEasanenrichedBODIPYbandismovedtotheleft,totheright,andbacktotheleftoftheBPEbychangingthetotalowrate.(c)Fluorescencemicrographsshowingthepositionoftheenrichedbandafter25,100,and205sfortheexperimentinpanelb.(d)MeasuredelectriceldprolesinthevicinityofthesecondaryBPEduringtheexperimentinpanelb.ThesecondaryBPEspanstheregionfrom0to360m.TheproleswererecordedwhentheenrichedbandresidedtotheleftofthesecondaryBPE(solidline),abovethesecondaryBPE(dashedline),andtotherightofthesecondaryBPE(dottedline).Theinitialconcentrationsinthemicrochannelwere5.0mMTrisHCl(pH8.1)and1.0.TheenrichmentandsecondaryBPEshadlengthsof500and360m,respectively.=25.0V. (Figure5d,dashedline)andtotherightofthesecondaryBPE(Figure5d,dottedline),theelectriceldstrengthovertheBPEincreasessignicantly.Thisincreasecorrespondstothemove-mentofthedepletionregionoverthesecondaryBPEsuchthatagreaterpotentialdropoccursinthisregion.Anelectricstrengthof8.0kV/mcorrespondstoa2.9VatthesecondaryBPE,whichissucienttoinducefaradaicreactionsatthesecondaryBPE.ThisresultdemonstratesthatthedepletionregioncanexertcontroloverthefaradaicreactionsatasecondaryBPE.Importantinformationaboutthelocationoftheenrichedbandcanbedeterminedfromsuchmeasurements.SUMMARYANDCONCLUSIONSWehavedescribedamethodformonitoringtheformationandpositionofanenrichedanionictracerbandinamicrochannel.cally,formationofanenrichedbandiscoincidentwithasharpincreasein,andthepositionoftheenrichedbandcorrelatestothemagnitudeof.Weusedbothexperimentsandsimulationstoexplaintheunderlyingmechanismforthisobservation.Lookingtothefuture,thendingsreportedhererepresentanimportantsteptowardourultimategoalofdevelopingalab-on-a-chipdevicecapableofsimultaneouslyenriching,separating,quantifyinganalytesonthemicroliterscaleandwithouttheneedforlabels.Havingeasyaccesstothelocationofenrichedbandsalsoprovidesasimplemeansforintegratingmorequantitativedetectionmethods,forexamplemassspectrometry,tothistypeofmicrodevice.Resultsrelatingtosuchstudieswill
8 bereportedinduecourse.ASSOCIATEDCONTENTS
bereportedinduecourse.ASSOCIATEDCONTENTSupportingInformation.Discussionofthefaradaicreac-tionsoccurringattheBPE,descriptionoftheemployednumer-icalapproachforsimulationofbuerandfaradaicreactions,anoteontheinuenceoftotalowrateontheformationofadepletionregion,anoteonthevariabilityofinuenceofPDFrateonenrichedbandposition,acontrolexperimentdemon-stratingtherelationshipbetweenenrichedbandconcentration,andanoteontheinuenceofBPElengthoninitiationoffaradaicreactions.ThismaterialisavailablefreeofchargeviatheInternetathttp://pubs.acs.org.AUTHORINFORMATIONCorrespondingAuthor*E-mail:crooks@cm.utexas.edu(R.M.C.);tallarek@stauni-marburg.de(U.T.).PresentAddressDepartmentofChemistry,UniversityofWashington,Box351700,Seattle,Washington98195-1700,UnitedStates.WegratefullyacknowledgesupportfromtheChemicalSciences,Geosciences,andBiosciencesDivision,OceofBasicEnergySciences,OceofScience,U.S.DepartmentofEnergy(contractno.DE-FG02-06ER15758).WealsothanktheRobertA.WelchFoundation(GrantF-0032)forsustainedsupport.SimulationswererunattheLeibniz-RechenzentrumderBayer-ischenAkademiederWissenschaften(Garching,Germany),supportedbyprojectHLRBpr26wo.MAJORSYMBOLSonsetpotential,Vpotentialdierencebetweentwoendsofthebipolarelectrode,Vappliedvoltagebetweenthedrivingelectrodes,VcurrentthroughtheBPE,Atotalcurrentthroughthemicrochannel,Atotalmicrochannellength,mtotalBPElength,mtotalsolutionresistanceinthemicrochannel,electroosmoticmobility,m/(Vs)electroosmoticvelocity,(1)Anand,R.K.;Sheridan,E.;Hlushkou,D.;Tallarek,U.;Crooks,R.M.LabChip,518(2)Anand,R.K.;Sheridan,E.;Knust,K.N.;Crooks,R.M.,2351(3)Perdue,R.K.;Laws,D.R.;Hlushkou,D.;Tallarek,U.;Crooks,R.M.Anal.Chem.,10149(4)Laws,D.R.;Hlushkou,D.;Perdue,R.K.;Tallarek,U.;Crooks,R.M.Anal.Chem.,8923(5)Hlushkou,D.;Perdue,R.K.;Dhopeshwarkar,R.;Crooks,R.M.;Tallarek,U.LabChip,1903(6)Dhopeshwarkar,R.;Hlushkou,D.;Nguyen,M.;Tallarek,U.;Crooks,R.M.J.Am.Chem.Soc.,10480(7)Ko,S.H.;Kim,S.J.;Cheow,L.F.;Li,L.D.;Kang,K.H.;Han,J.LabChip,1351(8)Mir,M.;Homs,A.;Samitier,J.(9)Salehi-Reyhani,A.;Kaplinsky,J.;Burgin,E.;Novakova,M.;Demello,A.J.;Templer,R.H.;Parker,P.;Neil,M.A.A.;Ces,O.;French,P.;Willison,K.R.;Klug,D.LabChip,1256(10)Du,Y.;Chen,C.G.;Zhou,M.;Dong,S.J.;Wang,E.K.,1523(11)Foote,R.S.;Khandurina,J.;Jacobson,S.C.;Ramsey,J.M.,57(12)Khandurina,J.;Jacobson,S.C.;Waters,L.C.;Foote,R.S.;Ramsey,J.M.Anal.Chem.,1815(13)Ramsey,J.D.;Collins,G.E.Anal.Chem.,6664(14)Chang,W.;Komazu,T.;Korenaga,T.Anal.Lett.,1468(15)Wen,J.;Wilker,E.W.;Yae,M.B.;Jensen,K.F.Anal.Chem.,1253(16)Macounova,K.;Cabrera,C.R.;Holl,M.R.;Yager,P.,3745(17)Hofmann,O.;Che,D.P.;Cruickshank,K.A.;Muller,U.R.Anal.Chem.,678(18)Tang,G.Y.;Yang,C.,1006(19)Ge,Z.W.;Wang,W.;Yang,C.LabChip,1396(20)Kelly,R.T.;Woolley,A.T.J.Sep.Sci.,1985(21)Burke,J.M.;Ivory,C.F.,893(22)Koegler,W.S.;Ivory,C.F.J.Chromatogr.,A,229(23)Petsev,D.N.;Lopez,G.P.;Ivory,C.F.;Sibbett,S.S.LabChip,587(24)Humble,P.H.;Kelly,R.T.;Woolley,A.T.;Tolley,H.D.;Lee,M.L.Anal.Chem.,5641(25)Shen,M.;Yang,H.;Sivagnanam,V.;Gijs,M.A.M.Anal.Chem.,9989(26)Chun,H.G.;Chung,T.D.;Ramsey,J.M.Anal.Chem.,6287(27)Wang,Y.C.;Stevens,A.L.;Han,J.Y.Anal.Chem.,4293 (28)McDonald,J.C.;Duy,D.C.;Anderson,J.R.;Chiu,D.T.;Wu,H.K.;Schueller,O.J.A.;Whitesides,G.M.,27(29)Hellmich,W.;Regtmeier,J.;Duong,T.T.;Ros,R.;Anselmetti,D.;Ros,A.,7551(30)Higuera,F.J.;Succi,S.;Benzi,R.Europhys.Lett.,345(31)Warren,P.B.Int.J.Mod.Phys.C,889(32)Capuani,F.;Pagonabarraga,I.;Frenkel,D.J.Chem.Phys.