Rapid measurement of phagocytosis by macrophages.

Publisher Website
Google Scholar
Abstract
A rapid method for measurement of phagocytosis by macrophages was investigated by the use of fluorescein-conjugated zymosan (Fl-zymosan) particles. Adherent peritoneal cells obtained from mice were used as macrophages. The fluorescence intensity of Fl-zymosan measured with a fluorescence spectrophotometer was proportional to the number of particles. Fl-zymosan particles, were incubated with macrophages in a flat-bottomed 96-well tissue culture plate at 37.degree.C in a CO2-incubator. After the removal a nonphagocytized particles, the fluorescence intensity of particles phagocytized by macrophages was measured with a fluorescence spectrophotometer (microplate reader). In the time course assay, the fluorescence intensity reached a maximum 20 min after the initiation of incubation, and thereafter maintained a constant level till 1 h in both normal and OK-432 injected groups. It was shown that the fluorescence intensity was dependent on the numbers of macrophages and Fl-zymosan particles in both normal and OK-432-injected groups when different numbers of particles (1 .times. 106, 5 .times. 106 or 1 .times. 107) were added to cultures each containing different numbers of macrophages (1 .times. 105, 2 .times. 105 or 4 .times. 105). The macrophages obtained from mice which were injected intraperitoneally (i.p.) with Propionibacterium acnes or proteose peptone as well as OK-432 showed higher phagocytic activity. Furthermore, it was also shown that the phagocytic activity measured with a fluorescence spectrophotometer agreed with that measured with a fluorescence microscope. The Fl-zymosan particles were applicable to the phagocytosis assay of opsonized particles. These results indicate that the method described here is useful for determination of macrophage phagocytic activity.