Advanced Photon Counting: Applications, Methods, Instrumentation

371 Pages·2015·13.05 MB·English

by  Peter Kapusta, Michael Wahl, Rainer Erdmann (eds.)

Springer Series on Fluorescence 15
Series Editor: Martin Hof
Peter Kapusta
Michael Wahl
Rainer Erdmann Editors
Advanced
Photon
Counting
Applications, Methods, Instrumentation
15
Springer Series on Fluorescence
Methods and Applications
Series Editor: Martin Hof
Springer Series on Fluorescence
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Advanced Photon Counting
Applications, Methods, Instrumentation
Volume Editors:
Peter Kapusta
Michael Wahl
Rainer Erdmann
With contributions by
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PeterKapusta MichaelWahl
J.HeyrovskyInstituteofPhysical PicoQuantGmbH
Chemistry Berlin
AcademyofSciencesoftheCzech Germany
Republic
Prague8
CzechRepublic
RainerErdmann
PicoQuantGmbH
Berlin
Germany
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SpringerSeriesonFluorescence
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Preface
Getherexcitedredorgreen;
letitdependonwhatshe’skeen.
Quitesoonquiteradiantshewillbeseen;
incolourstrueofhershe’llgleam,
butmindthenanosecondsinbetween.
Fortheymighttellwhichwaytolean:
perhapsshe’sotherthanyouhadforeseen.
Although,bepatientwhenyoutrythisscheme;
asingleflashthatlightsthescene
isnotenoughtogethertheme.
Infact,hermoodsaresoextreme
thattrulyrandomshemightseem.
Andyet,ifpatientlyyousiftherstream
apatternoftruebeautycanbeseen.
Sosumyourcountandbeserene.
In1926thephysicistFrithiofWolfersandthechemistGilbertN.Lewiscoinedthe
namephotonforthequantumoflightdiscoveredabout20yearsearlier.Evenifit
maylookalittlesuperficialatfirstglance,letusnotetheinvolvementofchemists
hereandelsewhereintheevolutionofquantumphysics.Indeed,theoverwhelming
successofquantummechanicsasamodernscientifictheoryisrootednotsomuch
inpurephysicsbutinitsinescapablyconvincingexplanatorypowerforvirtuallyall
aspectsofphysicalchemistryandmaterialscience.Modellingatomsandmolecules
as quantum mechanical systems undergoing transitions between quantum states,
some of them involving photon absorption and emission, was the key to under-
standing and eventually even exploiting virtually all previously mysterious spec-
troscopiceffects.Inthissenseitisnotasurprisethatthemethodsaddressedinthis
volume are now used more often in chemistry and related fields than in pure
physics.Infact,spectroscopicmethodshavebecomeindispensibleinbiochemistry
because light as a probe, suitably applied, can be used in living cells without any
damage to the specimen and without unduly spoiling the functions or processes
vii
viii Preface
under investigation. Even though classic spectroscopy does not require working
withsinglephotons,itturnsoutveryusefulthatitcanbedoneso.Thisisthecase
when there are only very few molecules involved in the processes of interest, in
particularinthevitalprocessesexploredbymolecularbiology.Evenmoreinterest-
ingthanclassicspectroscopyisthescenariowhereyetanotherquantummechanical
propertyisused:thelifetimeoftheexcitedstate.Itturnsoutthattheaveragetimea
moleculespendsinthisstateissospecifictothatmoleculeanditsenvironmentthat
itcanbeusedasafingerprintofthemoleculeinadditiontoitsspectrumand/orasa
probeforcertainenvironmentparameters.Eventhoughthelifetimeofanindividual
excited state is completely unpredictable according to quantum mechanics, the
averagelifetimeisbothmeasurableandmeaningful.Inpracticeitcanbeobserved
asthedurationofluminescencefromanensembleofmoleculesexcitedbyashort
flash of light. Going by the observed phenomenon one speaks of fluorescence or
phosphorescencelifetimemeasurements.Inthatcasethenecessaryaveragingofthe
individualexcitedstatelifetimesisachievedimplicitlybecauseofthesimultaneous
observationofphotonsfromtheentireensemble.Theotherinterestingscenariois
thatofaveragingtheexcitedstatelifetimeacrossmultiplecyclesofexcitationand
photonemissionobservedonjustonemolecule.Indeed,byvirtueofergodicity,this
kindofmeasurementgivesthesameresultsastheensemblemeasurement.Inthis
case time-correlated single photon counting (TCSPC) is the method of choice. It
allowsluminescencelifetimemeasurementsonsinglemoleculesandotherisolated
quantumsystems.Combining spectralinformationand,e.g.,fluorescencelifetime
onecanusethisrefined“fingerprint”ofthemoleculesofinteresttoidentifythem
even inthepresence ofsignificant background. Itwaslargelythisidea,combined
withconfocaldetection,whichfinallyledtotheincredibleachievementsinsingle
moleculedetection,singlemoleculespectroscopy,andevenmicroscopybymeans
of TCSPC. Powerful methods such as the exploitation of Fo¨rster resonant energy
transfer (FRET) asa molecularruler became routine toolsfor the investigationof
protein folding and interaction when they were made accessible for single mole-
culesalongthisway.
However,TCSPCisnotonlyusefulwithsinglemolecules.Aswewillshowin
thefirstchaptersofthepresentvolume,italsohelpstoachievebettertimeresolu-
tion with typical detectors, even in ensemble measurements. These technology
related chapters will cover the state of the art of current hardware, hopefully also
convincing the reader that despite the inherent statistical nature of the measure-
ment, modern users can rely on incredibly fast instruments and must not be as
patientasourlittleold-fashionedopeningpoemmightsuggest.Indeeditismostly
thememoryof“oldfashioned”instrumentationthatlefttheimpressionofslowness
attached to TCSPC. This may be illustrated by personal memories held probably
not only by the editors: “I recall with nostalgia the long nights spent alone in a
dimmedbasementlabwaitingforcollectionofatleast1000countsinthepeakby
our beautiful TCSPC monster powered by a 45 kHz flashlamp.” This is the past.
Meanwhile things have changed dramatically, not only because of faster TCSPC
electronicsbutverymuchalsobecauseoffaster(andeasiertouse)lasers,towhich
achapterinitsownwillbededicated.Theremainderofthechaptersarecoveringa
Preface ix
rich eclectic mixture of application topics as well as methodology in experiment
anddataanalysis.Despiteoftheimportanceoflifescienceapplicationswetriedto
embrace a much wider scope, including, e.g., defect centers in diamond as single
photon sources and quantum sensors, as well as optical tomography and super
resolutionmicroscopy.Similarly,onthemethodologyandinstrumentationside,we
aimedtoshowtheinterestingnewoptionsarisingfromthecombinationofappar-
ently distinct methods such as classic TCSPC and fluorescence lifetime with
methods based on intensity fluctuation. Together with the authors, to whom we
expressourgratitudehere,wehopetoprovideavolumeofbothanimmediatevalue
as a current overview of the field and some longer term value as a collection of
referencetexts.
Prague,CzechRepublic PeterKapusta
Berlin,Germany MichaelWahl
RainerErdmann