Molecular oxygen-sensitive fluorescent lipobeads for intracellular oxygen measurements in murine macrophages

Intracellular oxygen concentration is of primary importance in determining numerous physiological and pathological processes in biological systems. Th is paper describes the development and application of micrometer-sized oxyg en-sensitive fluorescence lipobeads for intracellular measurements of molec ular oxygen in J774 murine macrophages. A ruthenium diimine complex [Ru(bpy -pyr)(bpy)(2)]Cl-2 (bpy = 2,2 ' -bipyridine, bpy-pyr = 4-(q(1 " -pyrenyl)-2 ,2 ' -bipyridine) is used as the oxygen indicator. The indicator exhibits h igh chemical and photostability and high sensitivity to oxygen. The indicat or molecules are immobilized in a phospholipid membrane that coats polystyr ene microparticles. The fluorescence of the lipobeads is effectively quench ed by molecular oxygen. The fluorescence intensity of the oxygen-sensitive lipobeads is 3 times higher in a nitrogenated solution than in an oxygenate d solution. The lipobeads are internalized by murine macrophages through ph agocytosis. They maintain their spectral properties for 24 h in living cell s when the cells are stored in phosphate-buffered saline at pH 7.4. The pho tostability, reversibility, and effect of hypoxia, hyperoxia, and oxidative stress on the intracellular level of oxygen in J774 murine macrophages are described.