FLUORESCENT LIPOSOMES AS QUANTITATIVE MARKERS OF PHAGOCYTOSIS BY ALVEOLAR MACROPHAGES

A new phagocytic assay based on liposome ingestion by alveolar macrophages (AMs) is described. Fluorescent microspheres were encapsulated in liposomes, which allowed rapid enumeration by fluorometry. Liposomes made in the presence of vitronectin had the protein exposed on their outer surfaces, as determined by immunolabelling. Liposomes and rat AMs were incubated under conditions favorable for phagocytosis. Observation by light and electron microscopy showed AMs engulfing liposomes, with gradual transfer of fluorescent label from liposome to cell interior. This transfer was due to bona fide phagocytosis, as evidenced by (1) fluorescence of liposome-encapsulated dihydrofluorescein (DHF)-zymosan exclusively within AMs and (2) triggering of the respiratory burst by zymosan-associated liposomes only under conditions that allowed phagocytosis. Phagocytic activity was expressed as liposomes/cell, the average number of liposomes phagocytosed per macrophage. We used this technique to follow phagocytosis over time and to measure the effects of lipopolysaccharide (LPS) and vitronectin on AM phagocytosis.