5,10,15,20-tetrakis(N-methyl-4-pyridyl)-21H,23H-porphine (TMPyP) as a sensitizer for singlet oxygen imaging in cells: characterizing the irradiation-dependent behavior of TMPyP in a single cell.

Singlet molecular oxygen, a1Delta(g), can be detected from a single cell by its weak 1270 nm phosphorescence (a1Delta(g)-->X3Sigma(g)-) upon irradiation of the photosensitizer 5,10,15,20-tetrakis(N-methyl-4-pyridyl)-21H,23H-porphine (TMPyP) incorporated into the cell. The behavior of this sensitizer in a cell, and hence the behavior of the associated singlet oxygen phosphorescence signal, depends on the conditions under which the sample is exposed to light. Upon irradiation of a neuron freshly incubated with TMPyP, the intensity of TMPyP fluorescence initially increases and there is a concomitant increase in the singlet oxygen phosphorescence intensity from the cell. These results appear to reflect a photoinduced release of TMPyP bound to DNA in the nucleus of the cell, where TMPyP tends to localize, and the subsequent relocalization of TMPyP to a different microenvironment in the cell. Upon prolonged irradiation of the cell, TMPyP photobleaches and there is a corresponding decrease in the singlet oxygen phosphorescence intensity from the cell. The data reported herein provide insight into key factors that can influence photosensitized singlet oxygen experiments performed on biological samples.