The confocal laser scanning microscope has become a powerful tool in the study of the 3-dimensional distribution of proteins and specific nucleic acid sequences in cells and tissues. This is also proving to be true for a new generation of high contrast intermediate voltage electron microscopes (IVEM). Until recently, the number of labeling techniques that could be employed to allow examination of the same sample with both confocal and IVEM was rather limited. One method that can be used to take full advantage of these two technologies is fluorescence photooxidation. Specimens are labeled by a fluorescent dye and viewed with confocal microscopy followed by fluorescence photooxidation of diaminobenzidine (DAB). In this technique, a fluorescent dye is used to photooxidize DAB into an osmiophilic reaction product that can be subsequently visualized with the electron microscope. The precise reaction mechanism by which the photooxidation occurs is not known but evidence suggests that the radiationless transfer of energy from the excited-state dye molecule undergoing the phenomenon of intersystem crossing leads to the formation of reactive oxygen species such as singlet oxygen. It is this reactive oxygen that is likely crucial in the photooxidation of DAB.
Real‐Time In Vivo Detection of Cellular Senescence through the Controlled Release of the NIR Fluorescent Dye Nile Blue
