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June2008JournalofNortheastAgriculturalUniversity

    ;ApplicationandDevelopmentofBiosensors

    ;GAOYuelin,LIJichang,HUOGuicheng,andLIULibo

    ;CollegeofVeterinaryMedicine,NortheastAgriculturalUniversity,Harbin150030,China ;,,01.15No.28083

    ;Abstract:Asanewkindofanalyticalinstrument,theprinciples,typesandcharactersofbiosensorswerediscussedinthispaper.A

    ;biosensorisusuallycomposedofarecognitionelementofbiologicaloriginandaph)sicochemicaltransducer.Thebiologicalelement

    ;iscapableofsensingthepresence,activityorconcentrationofachemicalanalyseinsolution.Thesensingtakesplaceeitherasa

    ;bindingeventorabiocatalyticalevent.Theseinteractionsproduceameasurablechangeinasolutionproperty,inwhichthe~ansducer

    ;isconvenedintoaquantifiableelectricalsigna1.Theprinciples,typesandapplicationsofbiosensorsinenvironmentalinspection,food

    ;production,clinicalmedicineandmilitarydefensewerereviewed,andthetrendsinresearchwerepredicated.Furthermore,anattempt

    ;hadbeenmadetodescribethefuturedevelopmentdirectionsandprospects. ;Keywords:biosensors,development,application

    ;CLCnumber:TP212.3Documentcode:AArt:icleID:10068104(2008).0200800

    4

    ;Introduction

    ;Asanimmobilizationmatrixandapartofthesensing

    ;element,electrochemicallygrownpolymersarefind

    ;ingapplicationinchemicalandbiochemicalsensors.

    ;Conductingpolymerimmobilizationmatrixesarecon

    ;ceptuallyattractiveasameanofprovidingathree

    ;dimensionalconductingnetworkwherechargetrans

    ;fercanoccurbetweenaredoxspeciesandthepolymer

    ;withintheporesofthematrixJIThisphilosophypro

    ;motedthestudyofincorporationofredoxenzymes

    ;withinpolymers.Inthefieldofbiosensors,theelectro

    ;chemicalimmobilizationtechniqueisintroducedasan

    ;alternativewayforenzymeentrapmentinapolymer

    ;matrix.Thissimpletechniqueinvolvestheelectro

    ;synthesisofafilmontheelectrodesurface,starting

    ;fromasolutioncontainingthemonomerandtheenzy

    ;me.Thisfasttechniqueisaonestepprocess.How

    ;ever,adrawbackisenzymeleachingfromthefilm.

;Thecovalentattachmentoftheenzymeontothepoly

    ;mersurfaceprovidesasimplemethodtoassemblere

    ;producibleenzymeelectrodes.Differentcrosslinkers

    ;canbeused,glutaraldehydebeingthemostcommon.

    ;Duetoitsstabilityandinterestingredoxproperties4J,

    ;polyaniline(PAA)hasarousedconsiderableinterestas ;animmobilizationmatrix.PAApossessestworedox ;coupleswithinausefulpotentialrangeformanyen

    ;zymeredoxcofactorinteractions.PAAconductivity ;canbechangedbyvaryingthepotentialdifference ;acrossthefilmand,therefore,switchingbetweenthe ;differentoxidationstates.Glucoseoxidase(GOD)is ;immobilizedinaPAAlayerduringelectrodeposition ;fromphosphatebuffercontaininganilineandGOD, ;andusedtodetermineglucosebyconsumptionof ;oxygenat0.5VversusAg/AgC1.Thenextfrontier

    ;insensortechnologyisthedevelopmentofstructurally ;integrated,miniaturizedchemicalandbiologicalsen

    ;sor(micro)arraysforhighthroughoutmultianalyse ;environmentalandmedicalapplications.Structural ;Received10October2007

    ;SupportedbytheOpenFundoftheKeyLaboratoryDairyScienceProject,MinistryofEduc

    ationofChina(KLDS200608B)

    ;GAOYuelin(1982

    ),male,Master,engagedintheresearchofveterinarypharmacologyandtoxicology.

    ;Towhomcorrespondenceshouldbeaddressed.Email:lijichang828@sina.com ;E-mail:xuebaoenglish@neau.edu.on

    ;

    ;GAOYuelineta1.ApplicationandDevelopmentofBiosensors?8

    ;integrationofaphotoluminescence(PL)basedsensing ;elementwithcurrentlightsourcessuchaslasersand ;lampsisnotpossible,andintegrationwithinorganic ;lightemi~ingdiodesinvolvesintricatedesignandpro

    ;cedures.Incontrast,asensingelementcanbeinte

    ;gratedwithanorganiclightemittingdevice(OLED) ;lightsourceinastraightforward,flexible,anduniquely ;simpledesign.Suchintegrationisdemonstratedre

    ;centlyforagasphaseoxygensensor.

    ;EnzymeBiosensors

    ;AglucosebiosensorhasbeenpreparedbydopingRA ;withnegativelychargedGODandisusedtodetect ;enzymaticallyformedhydrogenperoxide.GODis ;attachedcovalentlytopoly(2-aminoaniline)filmvia ;chloranilasamediatorandbifunctionalreagent.This

    ;electrodeisstablebutnotselective.PAAischosen ;asamaterialforapotentiometricglucosebiosensor. ;PAAservesasapHsensitivematrixduetoitshigh ;slopeofpotentialversuspH/J.Glucosesensorsare

    ;preparedbyusingtwoGODimmobilizationmodes

    ;(electrochemicaldopingofpreviouslyformedconduc

    ;tivePAAandentrapmentduringelectrodeposition

    ;intoalayerofnonconductivePAA)onabaredand/or ;PrussianBluemodifiedplatinumelectrode.Theresult

    ;ingsensorissensitivetoglucosewhenoperatedina ;traditionalanodicmanneratanoperatingpotentialof ;0.6VinapH7.3solution.However,aconsiderably ;highbiasoriginatingfromascorbicacidandaceta

    ;minophenisobservedduetotherelativelyhighper- ;meabilityofthePAAlayerdepositedfromtheacidic ;solution.Gelatin(G)isawater-solubleproteinob

    ;tainedfrompartialhydrolysisofcollagen.Ithascar- ;boxyl,aminoandhydroxylfunctionalgroups.The ;mostcharacteristicpropertyofgelatinisitsabilityto ;formreversibleelasticgelswhenchilledinwaterand ;itscapabilitytoabsorbwaterupto600%700%ofits

    ;weightwhenallowedtoswellincoldwater.Gelatin ;hardenswhenreactedwithsomealdehydesandmetal ;salts[.

    ;Theglucoseoxidaseelectrodeisthemostapplied ;amperometricglucosesensorformeasurementscarried ;outinvivoandinvitro.GODisastableenzymewhose ;structurehasbeenelucidatedelsewhere.Ingeneral, ;theoxidationcurrentofH202isproportionaltothe ;glucoseconcentrationinthesolution.

    ;Immunobiosensors

    ;Apromisingtoolforthedetectionofheavymetalsis ;theuseofassaysbasedontheinhibitionofvarious ;enzymes.Ureasehasbeenthemostcommonlyused ;enzymeforthedeterminationofmetalions.Itisstable ;andinexpensive,andcanbeinhibitedbymanymetals ;suchasmercury,copper,silver,cadmium,lead,nickel, ;cobalt,manganeseandzinc[.

    ;Despitetheirhigh

    ;sensitivityandselectivitytowardsheavymetals,urease ;inhibitionassayshavescarcelybeenusedfortoxicity ;assessmentandidentiflcationduetoproblemswith ;measuringureaseinhibitionincomplexenvironmental ;samples[.

;Theimmobilizationofenzymesonscreen

    ;printedelectrodeshasbeenexploitedasatoolto ;producemetalionbiosensors.Enzymeimmobilization ;canovercomedifficultiessuchasinstabilitythatarises ;fromtheuseofenzymesinelectrocatalysisandalso ;helpinthefeasibilityofmultipleuseofthesameen

    ;zyme[“.Enzymeimmobilizationonthescreenprinted

     ;workingelectrodealsoallowsmassproductionofin

    ;expensive,simpleandversatilebiosensors.Enzymes ;havenormallybeenimmobilizedbyphysicaladsorp

    ;tion,covalentattachmentorpolymerizationentrap

    ;mentontotheelectrodesurface.However,adecrease ;inenzymeactivityisnormallyobservedafterimmobi

    ;lization,whichisattributedtoalowerflexibilityin ;theimmobilizedstate[.

    ;Flexibilityhasbeenshown

    ;tObecrucialindeterminingthelevelofactivityof ;manyenzymesandseemstopresentahigherdecrease ;incrosslinkingimmobilizationthangelentrapment. ;Flexibilityisalsoassociatedwithproteindenaturation ;inthatmoreflexiblestructuresaremorelikelytoex

    ;perienceirreversibleconformationalchangesresulting ;inenzymeinactivation[.Sinceimmobilizedurease ;hasbeenreportedtobelesssensitivetometalionin

    ;http://publish.neau.edu.cn

    ;

    ;?82?JournalofNortheastAgriculturalUnWerSityVD1.15No.22008

    ;hibition,andbecausetheanalyticalperformanceof ;thesensorcanbegreatlyaffectedbytheimmobiliza

    ;tionprocess,theimmobilizationmethodsmustbeopti

    ;mizedtomaximizeenzymestabilityandsensitivity, ;alongwithitsaffinityforboththesubstrateandin

    ;hibitort.

    ;Otherfactorssuchasthethicknessofthe

    ;enzymelayer,theenzymeloadingandtheconditions ;foroptimumenzymeactivitymustalsobeconsidered ;whenimmobilizationisperformed.

    ;OpticalBiosensor

    ;Thereisgrowinginterestintheuseofdiffraction.- ;basedsensorsfordiagnosticmeasurements.Recently, ;Gohea1.[16]describeanewdiffractionbasedmethod

    ;fordetectingandquantilyingbiologicalmolecules. ;Inthisapproach,biomoleculesarepatternedona ;surfacetocreateadiffractiongratingandthesurface

    ;isilluminatedtogenerateadiffractionpattern.When ;analyzemoleculesboundtothepatternedsurface,the ;intensityofthediffractionpatternincreases,enabling ;detectionandquantificationoftheanalyse.Thisnew

     ;approachtobiosensingisrapid,versatile,andselec

    ;tive,makingitwellsuitedforstudyingawiderangeof ;analyses.

    ;Intheory,anybiomoleculethatcanbepatterned ;onasurfacecanbeusedtodetectitscorresponding ;bindingpartner,suggestingusefulapplicationsfor ;detectionofsmalldrugmolecules,DNA,andother ;medicallyimportantmetabolites.Goheta1.[16]initially ;demonstratethistechniquebymeasuringthebinding ;ofantibodyconjugatedinthemouseIgG/antimouse

    ;IgGsystem.Itisalsoshownthatmultipleanalysesin ;amixedsolutioncouldbedetectedbycrosspatterning ;ofdifferentprobemolecules.Later,thedetectionlimit ;ofthesystemisimproved40foldbyusingagoldcon

    ;jugatedsecondaryantibody.Whileantibodyantigen

    ;bindingmakesforanusefultestcase,othersystems ;mustbeexploredinordertodiffraction-basedsen.’

    ;sorstoreachtheirfullpotentialasdiagnosticdevices. ;Anespeciallyusefulcapabilityforthesensorwould ;betheabilitytodetectenzymeactivityandtoquantify ;E-mail:xuebaoenglish@neau.edu.cn

    ;enzyme&inetics.

    ;CapacitiveBiosensor

    ;Thecapacitivebiosensorasanewtypeofbiosensor ;isofhighsensitivity,simplestructureandeasytoin

    ;tegrate.IthasbeenUsedfordirectdetectionofspecific ;antibodies,antigens,proteins,DNAfragmentsand ;heavymetalionswithouttheneedforalabe1.The ;applicationsofcapacitivebiosensorhavebeenintro

    ;duced[17].

    ;Someexistingproblemsandtheirpossible

    ;solutions,aswellasthefuturetrendsforthedevelop

    ;mentofcapacitivebiosensorarealsoproposed.Inor- ;dertodeterminethechlorogenicacid(CGA)intobacco ;fastandaccurately,amolecularlyimprintedpolymer ;piezoelectricquartzcrystalbulkacousticwavebio

    ;mimeticsensorusingCGAastemplatemoleculesis ;developedonthebasisofthesensitiveresponseof ;piezoelectricquartzcrystalandthespecificselectivity ;ofmolecularlyimprintedpolymer[18J.Themodifying

    ;functionofcoatinglayerontheresponse,theimprint- ;ingeffectandtheoptimaldetectingconditionsofthe ;sensorarealsodiscussed.Thelinearrangeofthesen

    ;sorforCGAis5.0x10—1.0xl0mol?L’..Therecoveryis

    ;96.7%105:0%.andRSDis3.7%.ThecontentsofCGA ;inseveraltobaccosamplesaredeterminedbythesen

    ;sorhorseradishperoxidase(HRP)whichisimmobiliz

     ;edonSiO2(about60nm)nanoparticles.Inthisexperi

    ;ment,HRPcanremainahigherbioactivitywiththe ;Michealis-Mentenconstantof7.8tool?L’[9].Bvmeans

    ;ofcatecholmediatorinthesolution,HRP/SiO2modi

    ;fledelectrodedisplaysanexcellentelectrocatalyti

    ;calresponselinearlytothereductionofH2O2inthe ;concentrationrange.Thetimeforsteady-statecurrent ;reaching95islessthan10s.Surfaceplasmonreso

    ;nance(SPR)biosensorshavebecomeaneffective ;instrumentofmeasuringbiomoleculespecificinterac

    ;tion.Ithasbeenadvancedrapidlythattheveterinary ;drugresiduesinanimaltissuesandbodyfluidare ;determinedbySPRbiosensor.Themethodisconven

    ;tionalandsensitive.ThecoNugatedreactionof ;drugandantibodycanbedetectedinrealtime.The ;

    ;GAOYuelinet,.ApplicationandDevelopmentofBiosensors?83? ;fundamentalsandcharacteristicsofthemethodin ;determinationofsomeveterinarydrugresidueswere ;describedbrieflyinthisarticle.

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