Studying phagocytosis by live-cell scintillation proximity assay.

Max F. Perutz Laboratories, Vienna, Austria.

Phagocytosis of microorganisms, senescent cells, apoptotic bodies, and effete tissue material is an important process in host defense and tissue homeostasis. A method is described to measure, in living macrophages, the kinetics of particle engulfment and lysosome/phagosome targeting. Plasma membranes or lysosomes are labeled with tritiated lipids, followed by exposure of cells to scintillant microbeads. Because of the short range of tritium beta-particles, geometric factors, and the confinement of lipids to membranes, scintillation can only be elicited by tracer molecules in membranes immediately vicinal to the scintillant. When the plasma membrane.is labeled with [(3)H]cholesterol, a signal is produced on bead-cell contact and engulfment and then reaches steady state within 45 min. When lysosomes are labeled with nonhydrolyzable [(3)H]cholesterol oleyl ether, scintillation requires intracellular lysosome/phagosome attachment or fusion, and steady state is attained only after several hours. The live-cell scintillation proximity approach is useful for examining the effects of pharmacological and genetic manipulations on particle uptake and on lysosome/phagosome targeting.

PMID: 18369943 [PubMed - indexed for MEDLINE]