Confocal laser scanning microscopy of urinary bladder after intravesical instillation of a fluorescent dye

Objectives. To assess the potential of confocal laser scanning microscopy for imaging of the urinary bladder after intravesical instillation of a fluorescent dye. Methods. The study was performed on the bladder of male Copenhagen rats. For confocal fluorescence microscopy (CFM), a standard confocal laser scanning microscope (Zeiss LSM 410) was used. Before measuring, the fluorescent marker SYTO 17 was instilled intravesically. After 2 hours of incubation, the rat was killed, the bladder excised and opened, and CFM was performed starting from the surface going through the urothelium and superficial layers of the lamina propria. Except for the opening incision, the bladder was left intact and no biopsies were taken. After imaging, the bladder was sent for conventional histologic studies. Results. CFM allows imaging of cellular details of the entire urothelium (superficial umbrella cells, intermediate, and basal urothelial cells) and superficial layers of the lamina propria. CFM images are close to those obtained by standard microscopy after conventional hematoxylin-eosin staining. Cell structure (eg, shape, size, chromatin texture, nucleoli, mitotic figures, nuclear/cytoplasmic ratio), as well as the structure of the connective tissue (eg, collagen fibers, blood vessels, erythrocytes), can be studied, allowing a standard histologic evaluation. Furthermore, in contrast to conventional histologic evaluation, CFM provides three-dimensional information and allows the study of intact tissue representing the true in vivo situation. Conclusions. CFM enables the study of the microscopic anatomy of bladder mucosa in its in vivo state. In combination with optical fiber bundles, endoscopic microscopy of the bladder may be possible in the future.