Figure 3.
Figure 3. Bacteriostasis parallels phagosomal acidification. (A) Growth capacity of MRSA cultured in medium with variable pH (n = 5). Data are mean + SEM. (B) Representative data from flow cytometric analysis of isolated neutrophils after incubation for 60 minutes with dual-labeled SA bioparticles and 40% serum in shaken suspension. Phagocytosing neutrophils were gated on the basis of AF647 positivity. (C) Isolated neutrophils were incubated in suspension with dual-labeled SA bioparticles in the presence of 40% serum and were placed under the microscope. Median pHrodo (left panel) and AF647 (right panel) fluorescence intensity of individual intra- and extracellular bioparticles was determined after 60 minutes using fluorescence microscopy (representative experiment; n = 5). Horizontal lines are medians ± SEM, and data were compared by using Mann-Whitney U tests. ****P < .0001. (D) Neutrophils were incubated with dual-labeled SA bioparticles in suspension in the presence of 40% serum in either normoxic or hypoxic conditions (n = 6). pHrodo (left panel) and AF647 (right panel) fluorescence of neutrophils was determined after 120 minutes by flow cytometry. Data are mean ± SEM. **P < .01; ***P < .001 as determined by Student t test for paired samples.

Bacteriostasis parallels phagosomal acidification. (A) Growth capacity of MRSA cultured in medium with variable pH (n = 5). Data are mean + SEM. (B) Representative data from flow cytometric analysis of isolated neutrophils after incubation for 60 minutes with dual-labeled SA bioparticles and 40% serum in shaken suspension. Phagocytosing neutrophils were gated on the basis of AF647 positivity. (C) Isolated neutrophils were incubated in suspension with dual-labeled SA bioparticles in the presence of 40% serum and were placed under the microscope. Median pHrodo (left panel) and AF647 (right panel) fluorescence intensity of individual intra- and extracellular bioparticles was determined after 60 minutes using fluorescence microscopy (representative experiment; n = 5). Horizontal lines are medians ± SEM, and data were compared by using Mann-Whitney U tests. ****P < .0001. (D) Neutrophils were incubated with dual-labeled SA bioparticles in suspension in the presence of 40% serum in either normoxic or hypoxic conditions (n = 6). pHrodo (left panel) and AF647 (right panel) fluorescence of neutrophils was determined after 120 minutes by flow cytometry. Data are mean ± SEM. **P < .01; ***P < .001 as determined by Student t test for paired samples.

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