Synthesis of ZnO Nanoparticles with Tunable Emission Colors and Their Cell Labeling Applications

DOI:10.1021/cm903869r

SynthesisofZnONanoparticleswithTunableEmissionColorsand

TheirCellLabelingApplications

XiaoshengTang,EugeneShiGuangChoo,LingLi,JunDing,andJunminXue*

DepartmentofMaterialsScienceandEngineering,NationalUniversityofSingapore,BlkE3A,04-10,

7EngineeringDrive1,Singapore117574

ReceivedDecember28,2009.RevisedManuscriptReceivedApril21,2010

ZnOnanoparticleshavebeenstudiedforpotentialcelllabelingapplicationsoverthepastseveralyears.However,littleprogresshasbeenmadebecauseofthelimitedemissioncolorandpoorwaterstabilityofZnOnanoparticles.Inthiswork,ZnOnanoparticleswithvariousemissioncolors,includingblue,green,yellow,andorange,weresynthesizedthroughanethanol-basedprecipitationmethod.TheemissioncoloroftheZnOnanoparticlescouldbetunedbyadjustingthepHvalueoftheprecipitationsolution.Theas-preparedZnOnanoparticleswerethenencapsulatedwithsilicatoformZnO@silicacoreshellnanoparticles,toimprovethewaterstabilityoftheZnOnanoparticles.ThevisibleemissionsoftheZnOnanoparticleswerewellretainedaftertheyhadbeencoatedwithsilicashells.TheresultantZnO@silicacoreshellnanoparticlesexhibitedlowcytotoxityandwerepromisingincelllabelingapplications.

1.Introduction

Fluorescentprobeshavebeenwidelyusedtostudycellularmorphology,behavior,andphysiologicalfunctions.Toachievehigh-qualityimagingandtrackingofbiologicalcells,theseprobesshouldbewater-dispersibleandbiocompatible,withahighluminescentefficiency.Inthepastcentury,organicdyesandfluorescentproteinswerethemajorfluor-escentprobesusedforbiologicalandbiomedicalresearch.1However,theseorganicprobesarehighlysusceptibletophotobleaching,sotheyarenotsuitableforlong-termcellimagingapplications.2Thisshortcominghasledtoastronginterestincolloidalfluorescentsemiconductorquantumdots(QDs),whichhaveshowntobepromisingforcellimagingsince1998.3ThetoxicityofQDshasbeenwidelystudied,butthemechanismsofthetoxicityarestillinquestion.ThereleaseofheavymetalionssuchasCd2þisfrequentlyproposedtoexplainthetoxicityofQDs,4butrecentstudies

*Towhomcorrespondenceshouldbeaddressed.Fax:þ6567763604.Telephone:þ65-65164655.E-mail:msexuejm@nus.edu.sg.

(1)Alivisatos,A.P.Science1996,271,933.

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Manna,U.;Yang,W.;Wickham,J.;Scher,E.;Kadavanich,A.;Alivisatos,A.P.Nature2000,404,59.(c)Han,M.;Gao,X.;Su,J.Z.;Nie,S.Nat.Biotechnol.2001,19,631.

(4)(a)Hines,M.A.;Guyot-Sionnest,P.J.Phys.Chem.1996,100,468.

(b)Wood,A.;Giersig,M.;Hilgendorff,M.;Vilas-Campos,A.;Liz-Marzan,L.M.;Mulvaney,P.Aust.J.Chem.2003,56,1051.(c)Abdullah,M.;Shibamoto,S.;Okuyama,K.Opt.Mater.2004,15,1751.(d)Wu,Y.L.;Lim,C.S.;Fu,S.;Tok,A.I.Y.;Lau,H.M.;Boey,F.Y.C.;Zeng,X.T.Nanotechnology2007,18,5604.(e)Jamieson,T.;Bakhshi,R.;Petrova,D.;Pocock,R.;Imani,M.;Seifalian,A.M.Biomaterials2007,28,4717.(f)Hardman,R.Environ.Sci.Technol.2008,42,6264.

(5)(a)Schneider,R.;Wolpert,C.;Guilloteau,H.;Balan,L.;Lambert,

J.;Merlin,C.Nanotechnology2009,20,225101.(b)Dumas,E.;Gao,C.;Suffern,D.;Bradforth,S.E.;Dimitrijevic,N.M.;Nadeau,J.L.Environ.Sci.Technol.2010,44,1464.

havealsodemonstratedthatthesurfacechemistryandproductionofreactiveoxygenspecies(ROS)fromQDsplayacrucialroleintheirtoxicity.5Besidesthetoxicityconcern,producingandemployingheavymetalion-basedcom-poundsareharmfultohumanhealthandposeapotentialenvironmentalhazard.Withregardtotheinherentproblemsofheavymetalion-basedQDs,thesearchfornewcellularprobeshasbecomeincreasinglyurgentandimportant.Recently,muchattentionhasbeenpaidtoanalternativesemiconductor,ZnO,forcelllabelingapplications.6Inprinciple,ZnOnanoparticlesdonothaveanyvisibleexci-tonicemissionastheirbandgapis3.4eV,7whichisbeyondthevisiblewavelengthrange.Interestingly,ithasbeenobservedthatthefreshlypreparedZnOnanoparticlesareabletoemitgreenlightunderUVexcitationandthusareinterestingforcelllabelingapplications.AlthoughmuchefforthasbeenspentindevelopingZnOintoanewcelllabelingprobe,thefactthatlittleprogresshasbeenmadetodatewasunexpected.Generally,therearetwomajorpro-blemswithZnOincelllabelingapplications.Thefirstproblemarisesfromtheirlimitedemissioncolor.ItisbelievedthatthevisiblegreenemissionofZnOnanoparticlesisattributedtotheirsurfacedefects;8however,thesesurfacedefectshavenotbeenwell-definedsofar,andthecorrelationbetweenthesurfacedefectsandthevisibleemissionisnotyet

(6)Xiong,H.;Wang,Z.;Liu,D.;Chen,J.;Wang,Y.;Xia,Y.Adv.

Funct.Mater.2005,15,1751.

(7)(a)Jin,J.;Soon,G.K.;Jung,H.Y.;Taeghwan,H.Adv.Mater.

2005,17,1873.(b)Chen,Y.;Myeongseob,K.;Lian,G.;Mathew,B.J.;Peng,X.J.Am.Chem.Soc.2005,127,13331.(c)Tamar,A.;Gong,Y.;Polking,M.;Yin,M.;Igor,K.;Gertrude,N.;Stephen,O.B.J.Phys.Chem.B2005,109,14314.(d)Narayanaswamy,A.;Xu,H.;Pradhan,N.;Kim,M.;Peng,X.J.Am.Chem.Soc.2006,128,10310.

(8)(a)Xiong,H.;Xu,Y.;Ren,Q.;Xia,Y.J.Am.Chem.Soc.2008,130,

7522.(b)Moussodia,R.O.;Balan,L.;Merlin,C.;Mustin,C.;Schneider,R.J.Mater.Chem.2010,20,1147.

r2010AmericanChemicalSocietyPublishedonWeb05/06/2010pubs.acs.org/cm

3384Chem.Mater.,Vol.22,No.11,2010clear.SeveralattemptshavebeenmadetotunetheemissioncolorofZnOnanoparticlesthroughsurfacemodifications.Forinstance,itwasreportedthatvariouscoloremissionsofZnOnanoparticlescouldbeachievedbysurfacemodifica-tionwithpolymers,8butthemechanismsofthevariouscoloremissionsthroughthesepolymersurfacemodificationsarefarfrombeingunderstood.Todate,tuningtheemissioncolorofZnOnanoparticlesremainschallengingbecauseofthelackofaknownexactmechanism.

Besidesthelimitedemissioncolor,ZnOshowspoorstabilityinwater,andthisposesthesecondseriousproblemwithZnOnanoparticlesascelllabelingprobes.Ithasbeenwelldocumentedthattheequilibriumcon-centrationofZn2þinwaterisquitehighoverawidepHrange.7Thisallowstheas-preparedZnOnanoparticlestobedissolvedinwatereasilyoverawidepHrange.ItispossibletostabilizeZnOnanoparticlesinwaterthroughsurfacemodifications,butthisisusuallyatthecostoflosingtheiroriginalvisiblegreenemission.Forexample,Liuetal.reportedthatZnOnanoparticlescouldbestabilizedbybeingcoatedwithdiethanolamine(DEA)andoleicacid(OA).9TheresultantZnOnanoparticlescoulddemonstratevisibleblueemissiononlyat430nm,whichisnotidealforbiologicallabelingbecausemostcellsandtissuesalsoappearblueunderUVlight.

AlthoughthereisstillmuchtounderstandaboutthemechanismsforvisibleemissionofZnOnanoparticles,numerousresearchreportshaverevealedthatthesurfaceoxygenvacanciesplayanimportantroleinthevisibleemissionofZnO.6,8,9Thesurfaceoxygenvacanciesarealsorelatedtothesurface-boundOH-andþHþgroups.Bymodifyingthesurface-boundOH-orHgroupsthroughpHvalueadjustment,wemaybeabletooptimizethesurfaceoxygenvacanciesandthentunetheemissioncolorofZnOnanoparticles.Onthebasisofthisunderstanding,aprecipitationprocesswasdesignedtosynthesizeZnOnanoparticleswithtunableemissioncolors.Withthismethod,nootheradditivessuchassurfactantsorpoly-merswereaddedexceptforzincacetateandlithiumhydroxide(LiOH).ZnOnanoparticleswerepreparedbyprecipitationofZn2þpurelywithLiOHinethanol.OurexperimentalresultsshowedthatZnOnanoparticleswithvariousemissioncolorsweresuccessfullycreatedviaadjustmentoftheconcentrationofLiOH.However,theas-precipitatedZnOnanoparticleswerenotstableinwaterforalongperiodoftime.TostabilizetheZnOnanoparticlesinwater,theas-precipitatedZnOnanopar-ticleswerethenencapsulatedwithsilicatoformZnO@silicacoreshellnanostructures.Silicawasselectedastheshellmaterialbecauseofitsbiocompatibility,watersta-bility,andrichsurfacechemistry.10TheobtainedZnO@silicacoreshellnanoparticlesexhibitedexcellentwaterstability,andthevisibleemissionsofZnOwereretained.

(9)Fu,Y.;Du,X.;Sergei,A.K.;Qiu,J.;Qin,W.;Li,R.;Sun,J.;Liu,J.

(10)J.(a)Am.Lu,Chem.Y.;Ying,Soc.Y.2007D.;,Mayers,129,16029.

B.T.;Xia,Y.N.NanoLett.2002,

2,183.(b)Deng,Y.H.;Wang,C.C.;Hu,J.H.;Yang,W.L.;Fu,S.K.ColloidsSurf.,A2005,252,87.(c)Li,L.;Choo,E.S.G.;Yi,J.B.;Ding,J.;Tang,X.S.;Xue,J.M.Chem.Mater.2008,20,6292.(d)Li,Z.Q.;Zhang,Y.Angew.Chem.,Int.Ed.2006,45,7732.

Tangetal.

ThecelllabelingapplicationsofthecreatedZnO@SiO2nanoparticlesarealsodemonstratedinthiswork.

2.ExperimentalSection

RawMaterials.Lithiumhydroxide(LiOH,99%pure,Sigma-Aldrich),sodiumhydroxide(NaOH,99%pure,Sigma-Aldrich),potassiumhydroxide(KOH,99%pure,Sigma-Aldrich),calciumhydroxide[Ca(OH)2,99%pure,Sigma-Aldrich],zincacetate(99%pure,Sigma-Aldrich),ethanol(absolute,Sigma-Aldrich),tetraethoxysilane(TEOS,99%pure,Sigma-Aldrich),3-[2-(amino-ethyl)aminopropyl]trimethoxysilane(APTS,99%pure,Sigma-Aldrich),dimethylsulfoxide(DMSO,99%pure,Sigma-Aldrich),andultrapurewater(18MΩcm)wereusedwithoutfurtherpurification.

SynthesisofZnONanoparticles.ZnOnanoparticleswerepreparedthroughaprecipitationmethodbyusingLiOHastheprecipitationagentinethanol.Ingeneral,0.2mmolofzincacetatewasdissolvedin20mLofabsoluteethanol.Themixturewasdissolvedcompletelybybeingstirredatroomtemperaturefor30min;36mgofLiOHwasdissolvedin20mLofabsoluteethanol.Thezincacetate/ethanolsolutionwasthenaddedtotheLiOH/ethanolsolution.ThepHvalueofthesolutionwasmeasuredtobe12.Afterreactionfor2h,thesolutionbecameturbid,indicatingZnOnanoparticleswereformed.ThepHvaluesofthesolutionsweretunedto10,8,and6when14,10,and5.5mgofLiOHwereadded,respectively.TheobtainedZnOnanoparticlesthatprecipitatedatdifferentpHvalueswerefirstwashedusingabsoluteethanoltoremoveunreactedpre-cursors,andthewashingprocesswasrepeatedthreetimes.Thepurifiednanoparticleswerethendispersedinabsoluteethanolforstorage.

SynthesisofZnO@SiO2.Twentymillilitersofanethanolsolutioncontaining0.25mmolofZnOnanoparticlesand0.06mLoftetraethoxysilane(TEOS)wereaddedto4mLofawatersolution,andthenthesolutionwastreatedwithultrasoundfor10min.Subsequently,5mgofammoniadissolvedin10mLofethanolwasaddedslowlytothesolutionwithcontinuousstirring.ThepHvalueofthesolutionwas8.Thesolutionwastransferredtoa100mLcone-shapedbeakerandstirredfor24hunderanitrogenatmosphere.Thesolutionwasthencentrifugedat3000rpmforthreecycles.Thepurifiedparticleswerethendispersedinabsoluteethanolforstorage.

SurfaceModificationofZnO@SiO2.FivemillilitersofZnO@SiO2nanoaprticlesinethanolwastaken,andanadditional25mLofethanolcontaining100μLof3-[2-(aminoethyl)aminopropyl]tri-methoxysilane(APTS)wasadded.Afterbeingstirredfor10h,thesolutiondescribedabovewasheatedtorefluxfor1h.ThenAPTS-modifiedZnO@SiO2nanoaprticleswerecollectedthroughcentri-fugation,washedwithethanolseveraltimes,andfinallydispersedinwater.

CellLabeling.NIH/3T3cellswereplacedina96-wellplateandincubatedat37°Cin5%CO2inair.After24h,10μLofZnO@SiO2nanoparticlesat10μg/mLwasinjectedintoeachwell.Theparticleswerethenincubatedfor24hatpH7.4.Afterincubation,thecellswereobservedusinganOlympusIx2-DSUdiskscanningconfocalmicroscope.

MTTAssayforCytotoxicityofZnO@SiO2Nanoparticles.TheMTTsolutionwasfilteredthrougha0.22μmfilterandstoredat4°C.NIH/3T3cellsweredividedintoa96-wellplateatadensityof4000cellsperwellandwereincubatedwithZnO@SiO2nanoparticlesatdifferentconcentrationsfor24h.Followingthis,10μLofMTTwasaddedtoeachwell.Afterincubationat37°Cfor∼2-3hin5%CO2inair,thecellculture

ArticleChem.Mater.,Vol.22,No.11,20103385

Figure1.TEMimagesofZnOnanoparticlesthatprecipitatedinethanolatpH(A)12,(B)10,(C)8,and(D)6.Ineachpanel,thetoprightinsetshowstheHRTEMimageofasinglenanocrystalandthebottomrightinsetthehistogramofsizedistribution.

mediumwasremovedwithaneedleandsyringe.ThewellswerewashedthreetimesusingPBS,andthen200μLofDMSOwasaddedtoeachwell.After∼2h,theplatewasanalyzedusingamicroplatereader.

Characterizations.Alltransmissionelectronmicroscopy(TEM)imageswereobtainedusingJEOL100CXinstrument(200kV).Wepreparedsamplesbydippingcarbon-coatedcoppergridsintothesamplesolutionandthendryingthematroomtemperature.Photoluminescence(PL)spectrawerere-cordedinopen-sized1cmpath-lengthquartzcuvettesusingaPerkin-ElmerLS55spectrofluorometer.UV-visabsorptionspectrawererecordedonaUV-1601spectrophotometer.

3.ResultsandDiscussion

ZnOnanoparticleswereprecipitatedinethanolatpH12,10,8,and6.ThepHvaluewasadjustedbymodifyingtheconcentrationsofLiOHadded.TheTEMimagesandsizedistributionhistogramsoftheresultantZnOnano-particlesarepresentedinFigure1.WhenthepHvalueis12,asshowninFigure1A,theobtainedZnOnanopar-ticleswerewelldispersedandtheirparticlesizerangedfrom2to5nmwithanaverageparticlesizeof3.3nm.LatticefringewasclearlyobservedintheHRTEMimageofasingleZnOnanoparticle,andthelatticespacewas

(insetofFigure1A),whichcorrespondstothedis-2.8A

tancebetweentwo(0002)planesofZnO.TheparticlesizeofZnOnanoparticleswasslightlyincreasedwhenthepH

valuewasdecreased.AsshowninFigure1B,theaveragesizeoftheprecipitatedZnOnanoparticlesatpH10was∼4.5nm.TheparticlesizeoftheprecipitatedZnOnanoparticleswascontinuouslyincreasedwhenthepHvaluewasfurtherdecreased,i.e.,5.5nmatpH8and6.5nmatpH6(Figure1C,D).TheresultantZnOnano-particlesatpH8and6werewell-crystallizedasindicatedbytheirrespectiveHRTEMimage.Inthisethanol-basedprecipitationmethod,itisclearthatpHvalueisakeyfactorincontrollingthesizeoftheZnOnanoparticles.AtahighpHvalue,ZnOnucleiwereformedfasterandinalargeramountthanthoseatalowpHvalue.ThisledtocompetitiongrowthofZnOnanoparticlesatahighpHvalueandthusresultedinthesmallerparticlesize.TheSAEDpatternsofZnOnanoparticlesthatprecipitatedatdifferentpHvaluesweredetermined,andtheresultsareshowninFigureS1oftheSupportingInformation.AllthediffractionringsintheSAEDpatternswereascribedtothelatticesofZnOnanocrystals.

Figure2Ashowsthephotoluminescence(PL)spectraoftheZnOnanoparticlesthatprecipitatedatpH12,10,8,and6,whichwerelabeledassamplesA-D,respectively.WeobservedthatthePLexcitationpeakofthesampleswasshiftedincreasinglyfrom309to356nmwhentheprecipitationpHvaluewasdecreasedfrom12to6.TheredshiftofthePLexcitationwithadecreaseinpHvaluecouldbeascribedtothequantumconfinementeffect.

3386Chem.Mater.,Vol.22,No.11,2010Figure2.(A)PhotoluminescentspectraoftheZnOnanoparticlesthatprecipitatedatpH(A)12,(B)10,(C)8,and(D)6.TheZnOnanoparticlesweredispersedinethanol.Theexcitationwavelengthswere309,325,328,and356nmforthenanoparticlespreparedatpH12,10,8,and6,respectively.(B)CorrespondingphotographsofZnOnanoparticlespre-paredatpH(1)12,(2)10,(3)8,and(4)6under365nmexcitation.(C)ζpotentialsofZnOnanoparticlesthatprecipitatedatdifferentpHvalues.

TheparticlesizeoftheZnOnanoparticlestendedtobelargerwithadecreasingpHvalueandthusledtoadecreaseinthebandgapenergyandanincreaseintheabsorptionwavelength.Thesimilarquantumconfine-menteffectsofZnOnanoparticleshavebeenreportedpreviously.6,8Moreinterestingly,ZnOnanoparticlespre-cipitatedatdifferentpHvaluespresentedvariousvisiblecoloremissionsinethanolundertheirrespectivepeakUVexcitations.Forinstance,whentheprecipitationpHvaluewas12,theZnOnanoparticlesexhibitedblueemis-sionat455nmunder309nmexcitation.Theemissioncolorcouldbetunedtogreen(494nm,under325nmexcitation),yellow(570nm,under328nmexcitation),ororange(590nm,under356nmexcitation)whentheprecipitationpHvalueswereadjustedto10,8,or6,respectively.ThePLquantumyields(QY)oftheZnOnanoaprticlesweremeasuredascomparedtothestan-dardrhodamine6Gethanolsolution(QY=95%).Thequantumyieldsoftheblue,green,yellow,andorangeemissionswere26.2,30.1,34,and19%,respectively,as

Tangetal.

showninFigureS2oftheSupportingInformation.ThequantumyieldsoftheZnOnanoparticlesobtainedinthisworkarecomparablewiththequantumyieldresultsreportedbyXiaetal.8aHowever,thequantumyieldsofthisworkarelowerthanthequantumyieldachievedinSchneider’swork.8bAsreportedinref8b,thequantumyieldofZnOnanoparticlescouldreach59%withpoly-mersurfacemodification.Figure2Bshowsaphotographtakenwithadigitalcamera,showingthattheobtainedZnOnanoparticlesweredispersedinethanolverywell.UponUV365nmillumination,variousvisiblecoloremissions,suchasblue,green,yellow,andorange,wereobservedfortheZnOnanoparticlesthatprecipitatedatpH12,10,8,and6,respectively.

ItappearedthatthevisibleemissionofZnOnanopar-ticlesalsounderwentaredshiftwithanincreaseinparticlesizeduetothequantumconfinementeffect.However,thisshouldnotbethecasehereasthevisibleemissionofZnOnanoparticlesisnotgeneratedfromthetransferofanelectronfromtheconductionbandtothevalenceband.AlthoughthemechanismsofZnOvisibleemissionarestillunderdebate,muchpreviousresearchsuggestedthatZnOvisibleemissionistheresultofthetransitionsinvolvingtrappedlevels,andthatsurfaceoxy-genvacanciesplayanimportantroleintheformationofthesetrappedlevels.6,8ThesurfacechargesoftheZnOnanoparticlesatdifferentpHvalueswerethenmeasured,andtheresultsarepresentedinFigure2C.WhenthepHwas12,thesurfacechargewas-30mV.WithadecreaseinthepHvalue,thesurfacechargebecamemorepositive,i.e.,-28mVatpH10,5mVatpH8,and15mVatpH6.

Zn2þ-OH-þOH-fZn2þ-O2-þH2O

ð1ÞZn2þ-OH-þHþfZn2þþH2O

ð2Þ

Thesurfacechargeofoxidesinsolutionisdeterminedbytheinteraction-betweenasurface-attachedOH-groupandOHorHþgroupsinthesurroundings,dependingontheisoelectricpointoftheoxides.WhenthepHvalueisabovetheisoelectricpoint,theresultantoxidesappeartohaveanegativechargebecauseoftheformationofO2-onthesurface(eq1),andviceversa,thesurfacechargeispositiveduetotheformationofZn2þbelowtheisoelectricpoint(eq2).FromthesurfacechargeresultsofZnOnanoparticlesshowninFigure2C,weestimatedthattheisoelectricpointofZnOnanoparticlesinthisworkwasbetween8and10.Fromtheanalysispresentedabove,wecouldinducethattheZnOsurfacesweredominatedbyO2-ions(lessoxygenvacancy)atpH12and10,whilethesurfacesweredomi-natedbyZn2þ(moreoxygenvacancy)atpH8and6.Therefore,wedeterminedthatthewavelengthofthevisibleemissionofZnOnanoparticleswasincreased(frombluetoorange)withanincreaseinsurfaceoxygenvacancy.Wehavetosaythatthisconclusionwasveryphenomenon-based,butitatleastprovidedusefulinformationforthemechanisticstudiesofZnOvisibleemission.WehavealsousedKOH,NaOH,andCa(OH)2toprecipitateZnO

ArticleFigure3.(A)TEMimageofZnO@silicacoreshellnanospheres.ZnOwaspreparedatpH10.Theinsetisthehigh-magnificationimageofasinglenanosphere.(B)TEMimageofZnO@silicacoreshellnanospheres.ZnOwaspreparedatpH8.Theinsetisthehigh-magnificationimageofasinglenanosphere.(C)TEMimageofZnO@silicacoreshellnanospheres.ZnOwaspreparedatpH6.Theinsetisthehigh-magnificationimageofasinglenanosphere.WepreparedalltheTEMsamplesoftheZnO@silicacoreshellnanoaprticlesbydippingcarbon-coatedcoppergridsintothesamplesolutionandthendryingthematroomtemperature.(D)Photo-luminescentspectraofthethreetypesofZnO@silicananospheresinwater(excitationwavelengthof365nm),showingthatthePLpropertiesoftheZnOnanoparticleswerewellretainedaftertheyhadbeencoatedwithsilica.

nanoparticlesinethanolsolutions.Wedemonstratedthatvariousvisiblecoloremissionscouldalsobeachievedbyadjustingtheirrespectiveconcentrations,asshowninFigureS3oftheSupportingInformation.

TheZnOnanoparticleswithvariousvisiblecoloremis-sionsarecertainlyinterestingforcelllabelingapplica-tions.However,theresultantZnOnanoaprticles,whichprecipitatedinethanol,werenotstableinwater.AsshowninFigureS4oftheSupportingInformation,visibleemissionsoftheZnOnanoparticlesdegradedfastwhentheparticlesweretransferredfromethanoltowater.Itwasbelievedthatthedegradationinvisibleemissionwasduetothecorrosionofthenanoparticlesurfaceinwater.Therefore,itisofutmostimportancetoprotectthenanoparticlesurfacebyformingachemicallystablecoat-ing.Thesurfacecoatingshouldalsoenhancethehydro-philicityandconfercolloidalstabilityontheZnOnanoparticlesinwater.Inthiswork,theZnOnanoparti-cles,whichwereprecipitatedatpH10,8,and6,werethenencapsulatedwithsilicashells.ThecorrespondingZnO@SiO2coreshellnanostructuresarepresentedinFigure3A-C,respectively.ToweakentheinfluenceoftheencapsulationprocessonsurfacestatesofZnOna-noparticles,theencapsulationprocesswasconductedinabsoluteethanolandatamildpHvalue.FromtheTEMimages,itwasobservedtherewasnoobviousdifferenceamongthethreeZnO@silicacoreshellnanostructures.TheaveragesizeoftheZnO@silicacoreshellnanostruc-tureswas∼50nm.Thehigh-magnificationTEMimageofasinglesphere(insetofeachTEMimage)clearlyshowsthatZnOnanoparticleshavebeensuccessfullyencapsu-latedwithsilicashells.Roughly,therewere∼20ZnO

Chem.Mater.,Vol.22,No.11,20103387

Figure4.(AandB)DICphotographandfluorescentimage,respectively,ofthecellslabeledwithZnO@silicananoparticleswithgreenemission.(CandD)DICphotographandfluorescentimage,respectively,ofthecellslabeledwithZnO@silicananoparticleswithyellowemissiom.(EandF)DICphotographandfluorescentimage,respectively,ofthecellslabeledwithZnO@silicananoparticleswithorangeemission.(G)CellviabilityofincubatedNIH/3T3cellswithincreasingconcentrationsofZnOnano-particlesfor24h.CellviabilitywasmeasuredusingtheMTTassay.

nanoparticlesineachsilicasphere.ThePLspectra(Figure3D)showthatthephotoluminescencepropertiesoftheZnOnanoparticleswerewellpreservedaftertheyhadbeencoatedwithasilicashell.ThequantumyieldsoftheZnO@silicacoreshellnanoaprticlesweremeasuredtobe18,20,and12%forthegreen,yellow,andorangeemissions,respectively.ThedecreaseinthequantumyieldsofthecoreshellnanoparticlesascomparedtothepureZnOnanoparticlescouldbeascribedtotheabsorp-tionofaphotonbythesilicashells.ToimprovethesuspensionstabilityoftheZnO@SiO2nanoparticlesin

3388Chem.Mater.,Vol.22,No.11,2010water,theobtainedcoreshellnanoparticleswerefurthermodifiedwith3-[2-(aminoethyl)aminopropyl]trimethoxy-silane(APTS)tointroducepositivelychargedNH2groups.Thesurfacechargeofthemodifiedcoreshellnanoparticleswasmeasuredtobe∼30mV.ThepositivelychargedZnO@silicananoparticlescouldbewelldispersedinwater,formingstablecolloidsolutions.ThewaterstabilityofthepositivelychargedZnO@SiO2nanoparticleswasfurtherstudiedunderdifferentaqueousconditionswithpHvaluesrangingfrom4.5to11.WefoundthattheZnO@SiO2nanoparticleswerestableinwatersolutionswhenthepHvaluesofthesolutionsrangedfrom4.5to9.WhenthepHvaluewasabove9,thepositivelychargedZnO@SiO2nanoaprticleswereaggregatedtogetherandsettledinseveralhours.ThestabilityofthepositivelychargedZnO@SiO2nanoparticlesinaPBSsolutionwasalsostudied.WedemonstratedthattheobtainednanoparticlescouldbewellsuspendedinPBSsolutionsformorethanoneweek,showingtheirgoodstabilityinPBSsolutions.Figure4showsthedifferentialinterferencecontrast(DIC)photographsandfluorescentimageofNIH/3T3cellsincubatedwithZnO@silicananoparticleswithdif-ferentemissionwavelengths.AsshowninFigure4B,thegreenemissionfromthecellswasclearlyobservedunderthefluorecentmicoscope,indicatingthatZnO@silicananoparticlesweresuccessfullyattachedtothecellsur-face.ThesuccessfulattachmentoftheZnO@SiO2na-noaprticlescouldbeascribedtotheirpositivesurfacecharges.Thecorrespondingcontrolexperiment,inwhichthecellswereincubatedwithoutadditionofZnO@silicananoparticles,isshowninFigureS5oftheSupportingInformation.Similarly,panelsDandFofFigure4showyellowandorangeemissions,respectively,withgood

Tangetal.

intensitysignals.Furthermore,thecytotoxicityoftheZnOnanoparticleswasmeasuredusingtheMTTassay,andtheresultsareshowninFigure4G.Wedemonstratedthat,afterincubationfor24h,morethan85%ofthecellssurvivedwhentheconcentrationofZnOnanoparticleswase30μg/mL,suggestingthatZnOnanoparticlesarequitesafetolivingcells.TheseresultsshowthatZnOnanoparticlesareverypromisingforapplicationsincelllabeling.

4.Conclusions

Insummary,ZnOnanoparticleswithvariousemissioncolorsofblue,green,yellow,andorangeweresuccessfullypreparedbyusingafacileethanol-basedprecipitationmethod.TheemissioncoloroftheZnOnanoparticleswasreadilytunedviaadjustmentofthepHoftheprecipita-tionsolution.Theas-synthesizedZnOnanoparticleswerenotstableinaqueoussolutions.TostabilizetheZnOnanoparticlesinwater,weencapsulatedtheas-precipi-tatedZnOnanoparticleswithsilicashellstoformZnO@-silicacoreshellnanostructures.TheZnO@silicacoreshellnanostructuresexhibitedgoodwaterstability,andthePLpropertiesoftheZnOnanoparticleswerewellretained.TheinvitrocellcultureexperimentsindicatedthattheZnO@silicacoreshellnanoparticlesexhibitedlowcytotoxicityandpromiseincelllabelingapplications.Acknowledgment.ThisworkwassupportedbytheSinagporeMOE’sARFTier1funding(WBSR-284-000-072-112).

SupportingInformationAvailable:Fiveadditionalfigures.ThismaterialisavailablefreeofchargeviatheInternetathttp://pubs.acs.org.