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SelectedArticleEnglishEditionNo.
13February201072SelectedArticleTheWorld'sMostSensitiveSpectrouorometersRayKaminsky,StephenM.
CohenHORIBAJobinYvonmanufacturesalargerangeoffluorescencespectrofluorometers,includingbenchtopmodelsthatmakesteady-statemeasurementtoadvancemodularsystemsthatcanalsomeasureuorescenceanisotropiesandlifetimesofyoursamples.
IntroductionByvirtueofitsemissionnature,fluorescenceisanextremelysensitivetechnique,ascompared,forexample,toinstrumentationinvolvingabsorption.
Youcanthinkofemissiontechniquesasanalogoustoweighingapennybyplacingitdirectlyonascale,whileabsorptiontechniquesarecomparabletoweighinganelephantwithandwithoutapennyonitsback,thensubtractingonevaluefromtheothertoobtaintheweightofthepenny.
Becauseoffluorescencesensitivity,instrumentstomeasuretheemittedlightfromtheUVthroughIRarewidelyemployedinareasofbiochemistryandtechnology,toprobemoleculardistancesaswellasthelocalenvironmentofstructuresandcells.
Theyarealsocommoninmaterialsscience,especiallynowthatnanotechnologyrisesinimportance.
Intheworldofsensitivity,theinstrumentsthatareunchallengedintheirabilitytodetecttheweakestofsignalsarethosefromHORIBAJobinYvon.
Wheneverwesetouttodesignanewgenerationofspectrofluorometers,ourfirstcriteriontoconsiderissensitivity.
Andsensitivityisachievedbyattentiontodetailssuchasoptics,electronics,andsoftware.
ABriefHistoryTheoriginalFluorolog,in1976,wasthemostsophisticatedspectrofluorometeravailable,distinguishedasthefirstcomputerizedfluorometer.
Itssuccessor,theFluorolog-2,appearedintheearly1980s,andthelatestgeneration,theFluorolog-3,wasfirstsoldin1996.
Ourfirstbenchtopmodel,theoriginalFluoroMax,wasintroducedin1990,anditsgreat-grandchild,theFluoroMax-4,enteredthemarketin2006.
Tocomplementoursteady-stateandfrequency-domainlifetimespectrofluorometry,HORIBAJobinYvonacquiredtheScottishfirmIBHin2003toincludetime-correlatedsingle-photon-countingtechnologyinoursystems.
HORIBAJobinYvon'smaincenterofresearchanddevelopmentislocatedinEdison,NewJersey.
Wecontinuetobetheleadingmanufacturerofthebestspectrofluorometricinstrumentsavailable,andboastthehighestsalesofanycompanyinthefield.
All-ReectiveOpticsOneofthereasonsourinstrumentsaresosensitiveisthefactthatinsteadoflenses,mirrorsareemployed.
Thebiggestadvantageofmirrorsistheirabilitytofocusorcollimatelightwithoutchromaticaberrations.
Thismeansthatlightisinfocusnomatterwhatwavelengthisinvolved.
Lensesareusuallyinfocusatonlyonewavelength,and,asyoumoveawayfromthefocus,theabilityoftheinstrumenttotransferandcollectlightislimited,especiallywhensources,samples,anddetectors,aresmall.
EnglishEditionNo.
13February2010TechnicalReports73PhotonCountingAnotherfeaturethatcontributestooutstandingsensitivityiscountingthepulsesinaphotomultipliertubethatresulteachtimeaphotonsstrikethesedetectors.
Instrumentsthatemployanalogtechniquessumtogethernotonlythecurrentoriginatingfromphotons,butalsothatduetonoiseintheelectronics.
Thislimits,ofcourse,theabilitytodiscriminatetherealsamplesignalfromthebackground.
Blazed,ruledgratings,selecteddetectors,focallengths,andsourcesalsocontributetothesensitivityoftheseinstruments.
Sowhydon'tothersystemsincorporateallthesefeaturesaswellCostistheprimaryreason.
Ifyouwantsensitivity,youhavetopayforit.
WhoNeedsSensitivitySensitivityismorethantheabilitytodetectsignalsfromsamplesatlowerconcentrations.
Ifyoucountmorephotons,obviouslytheaccuracyofyouranswersismuchhigherasyourstatisticalsampleisgreateraswell.
Inaddition,moresignalmeansyoucantakedatamuchfasterandachievethesameaccuracy.
Sensitivity,accuracy,andspeed.
That'swhatyougetwithinstrumentsfromHORIBAJobinYvon.
Bench-topSimplicityorModularFlexibilityTomeetthedemandsofresearchersinmanyfields,HORIBAJobinYvonofferstwotypesofinstruments:theFluoroMax-4andtheFluorolog-3.
TheFluoroMax-4(Figure1),thenewestspectrofluorometer,isabenchtopunitthatoffersallthesensitivityandaccessoriesnecessaryforstate-of-the-artfluorescenceinvestigations,butoffersthesimplicityandconvenienceofaself-containedunit.
Featureslikeself-calibratingwavelengths,computercontrolledslits,andautomatedexperimentsinsoftwarearestandard.
OrderaFluoroMax-4anditarrivesreadytotakedatarightoutofthebox.
TheFluorolog-3(Figure2)isthemostadvancedinstrumentavailable.
Themostprominentfeatureismodularity,whichencouragesyoutodesignaninstrumentthatisidealforyouruniqueapplication.
Forexample,youcanadddoublemonochromatorstoachievehigherthroughputandbetterrejectionofbackgroundradiation,oryoucanaddaspectrographandCCDdetectortoobtaincompletespectrainamatterofmilliseconds(Figure3).
Youcanoperateintheultraviolet,visible,orinfraredbyselectingfromavarietyofgratingsanddetectors.
Youcanalsochoosetomountmultipledetectorsorsourcesthatcomplementyourapplication.
Figure2ModularSpectrouorometerFluorolog-3Figure3Near-IRExcitation/EmissionMatrixFullyCharacterizesThisNanoSampleOnaNanolog,ASpecialVersionoftheModularFluorolog-3Inadditiontoacquiringsteady-statespectraofsamples,accessoriescanbeaddedtobothFluoroMaxandFigure1TheFluoroMax-4Bench-topSpectrouorometerSelectedArticleEnglishEditionNo.
13February201074FeatureArticleTheWorld'sMostSensitiveSpectrouorometersFluorologtoacquirelifetimedata.
Fluorescencelifetimesaretheaveragetimebetweenabsorptionoftheexcitationlightandemissionoffluorescence.
Byacquiringlifetimedata,informationonthedynamicsofprocesseswithinthesampleswilltellyoumoreabouttheenvironment,size,anddistances.
HORIBAJobinYvonistheonlycompanythatsuppliesthetwomostprominentmethodsforacquiringlifetimedata:Time-CorrelatedSinglePhotonCounting(TCSPC(Figure4))forultimatesensitivity,orthephase/modulationmethodforthefastestacquisition,asfastasonemillisecond.
Thesetechniquescancharacterizelifetimesassmallasseveralpicoseconds.
Figure4TheTCSPCLifetimeOptionCanCharacterizeLifetimesasShortasPicosecondsConclusionThisgivesyouabriefintroductiontoHORIBAJobinYvon'sFluorescenceproductline.
Ofcourse,anyoftheseinstrumentscanbefittedwithavastvarietyofaccessories,fromheater/coolers,tomicroscopes,orpolarizerstodetermineanisotropy.
EnglishEditionNo.
13February2010TechnicalReports75RayKaminskyHORIBAJobinYvon,Inc.
MolecularandMicroanalysisDivisionFluorescenceSpectroscopyVicePresidentStephenM.
CohenHORIBAJobinYvonIncSeniorTechnicalWriterPh.
D.