CHAPTER 11: Imaging of Tissue Oxygen Ex vivo

In the last decade considerable progress has been achieved in the design of various fluorescent and phosphorescent oxygen sensing probes, their delivery to live animal tissue and subsequent imaging of tissue oxygenation. A significant number of publications reflect the growing interests in O₂ imaging technologies. The in vivo O₂ imaging platforms, traditionally more attractive, have limited applicability, e.g. to skin, thin layers of brain, muscle and some other tissues and organs, small animals such as zebrafish and a few other organisms. However, the range of applications can be significantly broadened by ex vivo tissue models: by using them, O₂ distribution and consumption rate can be conveniently analysed at a single cell level and cross-correlated with other functional parameters. Here we describe several high-resolution tissue O₂ imaging studies with ex vivo models using cell-penetrating phosphorescent O₂-sensitive probes. Detailed procedures for ex vivo fluorescence/phosphorescence lifetime imaging (FLIM/PLIM) of tissues and general algorithms of acquisition, processing and interpretation of the imaging data are provided, and the merits and limitations of this approach are discussed.