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Computer Image Analysis Method for Rapid Quantitation of Macrophage Phagocytosis
Author(s) -
Odeyale Charles O.,
Hook Gregory R.
Publication year - 1990
Publication title -
journal of leukocyte biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.819
H-Index - 191
eISSN - 1938-3673
pISSN - 0741-5400
DOI - 10.1002/jlb.48.5.403
Subject(s) - phagocytosis , macrophage , flow cytometry , biology , microsphere , cell , cell counting , microbiology and biotechnology , cell culture , biophysics , immunology , biomedical engineering , in vitro , biochemistry , cell cycle , medicine , genetics , chemical engineering , engineering
A new method is described for rapidly quantitating phagocytosis by adherent macrophages in culture using computer image analysis (CIA) of video light microscopic images. Ingestion of fluorescent microspheres by peritoneal murine macrophages is used to model phagocytosis. The grey levels of digital phase contrast and fluorescent microscopic images are used to quantitate the number of microspheres per cell. The method is semi‐automatic, analyzes approximately 2 × 10 3 cells/hr, and simultaneously measures phagocytosis (microspheres/cell), cell area, and density (number of cells/mm 2 ). CIA obtains the same microspheres/cell average as manual microscopic counting and an analytical precision of 5%. As expected, CIA found that the number of microspheres/cell linearly increases with increasing macrophage‐microsphere co‐culture time or increasing microsphere concentration until macrophages become saturated. CIA finds increased phagocytosis by interferon‐gamma‐treated cells and suppressed phagocytosis by cytochalasin B‐ or 4°‐treated cells relative to controls, which demonstrates that CIA can resolve biological changes in macrophage phagocytosis. CIA also provides quantitative data on macrophage morphometry and density and found an increase in the cell area and density of INF treated macrophages. CIA provides significantly more phagocytic, morphometric, and density data than conventional manual microscopic counting methods or flow cytometric methods. The limitations, improvements, and future applications of this method are discussed.