Laser line-scanning confocal fluorescence imaging of the photodynamic action of aluminum and zinc phthalocyanines in V79-4 Chinese hamster fibroblasts.

Confocal fluorescence microscopy, using a newly constructed laser line-scanning confocal microscope, was applied to an investigation of the early stages of photoinduced destruction of V79-4 Chinese hamster fibroblasts using aluminum and zinc phthalocyanines as photosensitizers. Results obtained in this work show that aluminum and zinc phthalocyanines, once internalized, localize in perinuclear sites that are disrupted upon light exposure resulting in fluorescence redistribution. The combination of laser-line scanning with charge-coupled device detection used in the confocal microscope developed in this work can enable rapid high-resolution sequential imaging, which is ideal for studying photoinduced intracellular fluorescence dynamics.

Development of a laser-based fluorescence microscope with subnanosecond time resolution.

This report describes the development of a fluorescence microscope based on a standard inverted optical microscope which incorporates a pulsed picosecond dye laser excitation source and a detector consisting of a gated image intensifier coupled to a CCD camera. Fluorescence images have been obtained using gate durations of 0.5 ns from this apparatus, representing a reduction in gate duration of an order of magnitude compared with similar instruments reported by others recently. Subnanosecond gated fluorescence images of V79-4 Chinese hamster lung fibroblasts stained with a phthalocyanine photosensitizer used in photodynamic therapy are presented. The results of these measurements are discussed in terms of the intracellular distribution of the sensitizer. Other potential applications and limitations of this technique are also outlined.