Fluorescence

Quantum Efficiency• Only emitted light is relevant for fluorescence detection in microscopy –intersystem conversion processes equals to loss of fluorescence efficiency• Quantenausbeute (quantum yield or quantum efficiency [QE]) in steady state:Number of emitted photonsQE = --------------------------------------Number of absorbed photons• QE is the essential for a fluorophore to qualifyfor optimal use in microscopyA good fluorophore can be excited up to 10 6 timesbefore it is „dead“Quantum Yield[Q.Y.]StandardsQ.Y. [%]Conditions forQ.Y.MeasurementsExcitation [nm]Cy3 4 PBS 540 2Cy5 27 PBS 620 2CresylVioletFluorescein5395POPOP 97QuininesulfateRhodamine 101Rhodamine 6GRhodamine BTryptophanL-Tyrosine58Methanol0.1 MNaOH,22 o CCyclohexane0.1 MH 2 SO 4 ,22 o C580 3496 3300 3350 3100 Ethanol 450 495 Water 488 431 Water 514 413Water,20 o C280 314 Water 275 3Ref.