FLUORESCENCE-LIFETIME IMAGING MICROSCOPY METHOD HAVING TIME-CORRELATED SINGLE-PHOTON COUNTING

摘要

The invention relates to a fluorescence-lifetime microscopy method having time-correlated single-photon counting, wherein a sample (13) is periodically excited by means of excitation light pulses to emit fluorescence photons using a pulsed light source (2), wherein a measurement interval is defined between each pair of consecutive excitation light pulses, the fluorescence photons are detected by means of a detector (16) and a detector signal (17) representing the detected fluorescence photons is produced, detection times at which the fluorescence photons are detected by the detector (16) within the measurement intervals are determined on the basis of the detector signal (17), and imaging is performed on the basis of the detection times. The aim of the invention is to increase the excitation light intensity with comparatively little technical effort whilst avoiding a pile-up effect and for universal use of detector and electronics types This aim is achieved in that, in the fluorescence-lifetime microscopy method, it is determined in the respective measurement interval whether a predetermined number of fluorescence photons has been detected within the measurement interval. The detection times for all the detected photons are aggregated in a first data memory, common to a plurality of image points. The detection times of only those fluorescence photons which have been detected in the predetermined number in the respective measurement intervals are aggregated in a second data memory, common to this plurality of image points. In a computational step, the detection times aggregated in the first data memory are combined with the detection times aggregated in the second data memory. The results of this computational step are stored in a third data memory. The invention further relates to a microscope for carrying out such a fluorescence-lifetime microscopy method.