Fluorescence methods for monitoring phagosome-lysosome fusion in human macrophages

Publisher Summary This chapter focuses on the fluorescence methods for monitoring phagosome- lysosome fusion in human macrophages.This chapter presents an alternative assay based on the use of sulforhodamine or rhodamine (R)-labeled dextran. The assay is based on the initial uptake of sulforhodamine or R-dextran into secondary lysosomes of macrophages, and the subsequent colocalization of the rhodamine label and phagocytosed fluorescein-labeled yeast cells on the fusion of phagosomes with secondary lysosomes. The macrophage pathogen Mycobacterium avium is thought to persist in phagosomes by preventing the fusion of the phagosomes with lysosomes. These observations support the hypothesis that live M, avium inhibits phagosomelysosome fusion, and that M-CSF can activate macrophages to overcome the inhibition of phagosome-lysosome fusion. The molecular mechanisms of the inhibition of fusion, and of the reversal of this process by macrophage colony-stimulating factor(M-CSF), remain to be investigated.