CHAPTER 13: Probing Tissue Oxygenation by Delayed Fluorescence of Protoporphyrin IX

Oxygen-dependent quenching of delayed fluorescence of protoporphyrin IX (PpIX) has been introduced a decade ago as a method for measuring oxygen at the mitochondrial level in cells and tissues. The technique makes use of the optical properties of 5-aminolevulinic acid-enhanced mitochondrial PpIX to measure mitochondrial oxygen tension (mitoPO2). Like phosphorescence quenching, the technique is very scalable and mitoPO2 measurements can be performed in isolated cells, isolated organs, in vivo and nowadays even in humans at the bedside in a clinical setting. Administration of aminolevulinic acid (ALA) induces measurable levels of PpIX in the mitochondria where PpIX acts as an oxygen-sensitive dye. PpIX emits a red delayed fluorescence after excitation with a pulse of green light. The lifetime of the delayed fluorescence is inversely related to mitoPO2. MitoPO2 can be calculated from the lifetime according to the Stern-Volmer equation. Dynamic mitoPO2 measurements during local cessation of oxygen supply allow for assessing oxygen disappearance rate as a measure for cellular respiration in vivo. In this chapter we will discuss the background of the technique and its implementation in both a laboratory and the clinical setting and provide examples of its application in experimental animals and man.

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