CONFOCAL SPECTRAL IMAGING ANALYSIS IN STUDIES OF THE SPATIAL-DISTRIBUTION OF ANTITUMOR DRUGS WITHIN LIVING CANCER-CELLS

Advanced instrumentation allowing the recording of confocal microfluor escence and micro-Raman spectra, with spatial resolution on the sub-mi crometre scale, has been developed. Computer-controlled automatic scan ning stage and a system of scanning the laser beam through the stage b oth permit the recording of well resolved spectral images of samples. Conventional images of species could be recorded with a CCD camera thr ough the same optics. The developed software includes the possibilitie s of converting a spectral image into a conventional image and of assi gning point-by-point the spectral data to the conventional image. This technique was applied to the analysis of the microfluorescent signals from living cancer cells treated with the antitumour drugs doxorubici n and mitoxantrone. It was possible to construct separately the spectr al images corresponding to the distribution of drug-DNA complexes, the free drug and intrinsic signals from untreated cell. Consequently, qu antitative comparative intracellular and even intranuclear localizatio n of antitumour drugs was achieved. The differences in the intracellul ar distribution of the drug-DNA complexes inside the cells are discuss ed in terms of biological effects.