SIMULTANEOUS CONFOCAL LIFETIME IMAGING OF MULTIPLE FLUOROPHORES USINGTHE INTENSITY-MODULATED MULTIPLE-WAVELENGTH SCANNING (IMS) TECHNIQUE

We demonstrate the simultaneous recording of confocal lifetime images of multiple fluorophores, The confocal microscope used in the study co mbines intensity-modulated laser illumination, lock-in detection and s pectral separation of the fluorescent light. A theoretical investigati on is presented that describes how the signal-to-noise ratio (SNR) dep ends on various factors such as modulation frequency, degree of modula tion and number of detected photons. Theory predicts that, compared wi th ordinary intensity images, lifetime images will have a SNR that is, at best, approximately four times lower. Experimental results are pre sented that confirm this prediction.