title: Dynamic quantitative assays of phagosomal function.
abstract: Much of the activity of the macrophage as an effector cell is performed within its phagocytic compartment. This ranges from the degradation of tissue debris as part of its homeostatic function to the generation of the superoxide burst as part of its microbicidal response to infection. We have developed a range of real-time readouts of phagosomal function that enable these activities to be rigorously quantified. This unit contains descriptions of several of these assays assessed by different methods of quantitation, including a fluorescence resonance emission transfer (FRET) assay for phagosome/lysosome fusion measured by spectrofluorometry, a fluorogenic assay for the superoxide burst measured by flow cytometry, and a fluorogenic assay for bulk proteolysis measured by confocal microscopy. These assays illustrate both the range of parameters that can be quantified and the flexibility of instrumentation that can be exploited for their quantitation.
authors: Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, New York.
authors: Maria
authors: M
authors: Podinovskaia
authors: Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, New York.
authors: Brian C
authors: BC
authors: VanderVen
authors: Department of Comparative Biology and Experimental Medicine, Faculty of Veterinary Medicine, University of Calgary, Calgary, Alberta, Canada.
authors: Robin M
authors: RM
authors: Yates
authors: Liverpool School of Tropical Medicine, Liverpool, United Kingdom.
authors: Sarah
authors: S
authors: Glennie
authors: Mid-Cheshire NHS Foundation Trust, Crewe, United Kingdom.
authors: Duncan
authors: D
authors: Fullerton
authors: Malawi-Liverpool-Welcome Trust Laboratories, Queen Elizabeth Hospital, Chichiri, Blantyre, Malawi.
authors: Henry C
authors: HC
authors: Mwandumba
authors: Department of Microbiology and Immunology, College of Veterinary Medicine, Cornell University, Ithaca, New York.
authors: David G
authors: DG
authors: Russell
mesh_terms: D000818:Animals; D002469:Cell Separation; D002478:Cells, Cultured; D005434:Flow Cytometry; D031541:Fluorescence Resonance Energy Transfer; D008264:Macrophages / Q000276:immunology*; D051379:Mice; D018613:Microscopy, Confocal; D010587:Phagocytosis; D010588:Phagosomes / Q000276:immunology / Q000378:metabolism*; D059748:Proteolysis; D013481:Superoxides / Q000378:metabolism
chemical_list: D013481:Superoxides
keywords: macrophage; phagocytosis; phagosome; phagosome maturation