The potential for the application of fluorescence lifetime imaging (FLIM) microscopy to studies of photosensitization mechanisms in photodynamic therapy (PDT) has been investigated. The fluorescence microscope incorporates a standard inverted optical microscope, a picosecond pulsed dye-laser excitation source, and an intensified CCD camera detector capable of being gated on a sub-nanosecond timescale. Fluorescence lifetime images resulting from multi-component analysis of sub-nanosecond gated fluorescence images of monolayer V79-4 Chinese hamster lung fibroblasts stained with disulphonated aluminium phthalocyanine (AlPcS2), a photosensitizer used in PDT, are presented. The results of these measurements are discussed in terms of the intracellular localization of the sensitizer. Preliminary results from multi-component FLIM of V79-4 cells multiply stained with AlPcS2 and a potential intracellular pH lifetime probe, 5(+6)-carboxynaphthofluorescein, are also presented.