Time-resolved fluorescence spectroscopy and intracellular imaging of disulphonated aluminium phthalocyanine

Spectroscopic studies were carried out on the photosensitizer disulphonated aluminium phthalocyanine (AlS₂Pc) which has prospective applications in photodynamic therapy. The fluorescence lifetimes of AlS₂Pc were measured in a range of model systems and cultured leukaemic cells using laser excitation and time-correlated, single-photon-counting detection. In an investigation of non-covalent protein binding, we studied AlS₂Pc in the presence of human serum albumin (HSA) in 0.1 M phosphate-buffered saline at pH 7.4. On addition of excess concentrations of HSA, small red shifts in the fluorescence and absorbance spectra were observed, together with an increase in fluorescence polarization anisotropy, consistent with binding of the phthalocyanine. Fluorescence decays could be resolved into two lifetimes for bound AlS₂Pc with a dominant component of 5.5 ns and a minor component of 1 ns. Fluorescence imaging and time-resolved microfluorometry were carried out on intracellular AlS₂Pc using leukaemic K562 cells. Microscopic imaging with a charge-coupled device (CCD) camera revealed that AlS₂Pc fluorescence predominated in a discrete perinuclear region which was then probed selectively by a focused laser spot for fluorescence lifetime measurements. Bi-exponential decays with lifetime components of 6.1 and 2.2 ns were observed. On irradiation at 633 nm, the fluorescence intensity increased initially and subsequently declined due to photodegradation.