The internalization of chitosan by PMNs. Freshly isolated polymorphonuclear neutrophils (PMNs) were resuspended in RPMI 1640 supplemented with 0.1% decomplemented fetal bovine serum, pre-stained with 1 μg/ml calcein-AM for 30 minutes at 37°C and incubated with 100 μg/1 × 106 cells rhodamine B isothiocyanate (RITC)-zymosan for 1.5 hours (a positive control), 15 μg/ml RITC-80% deacetylated (80 M) or RITC-95% deacetylated (95 M) chitosan for 3 hours at 37°C. PMNs were then centrifuged and plated on a slide coated with 100% decomplemented autologous serum and visualized by live confocal microscopy. The index of internalization of chitosan by PMNs was calculated as the percentage of cells that internalized RITC-chitosan. Results are presented as mean ± standard error. This figure represents the results of three independent experiments.