Effects of AT on purified human neutrophil morphology. (A-B,F-I) Purified human neutrophils were labeled with calcein-AM (green) and Hoechst33342 (blue) for 15 minutes. The neutrophils were incubated with different concentrations of HSA, HRG, AT, rAT, ARG, or HBSS at 37°C for 60 minutes on polystylene surface. The shape of neutrophils was observed under an IN Cell Analyzer, which determined the form factor (maximum diameter/minimum diameter) and average area of neutrophils. ∗∗∗P < .001 vs HBSS, †††P < .001 vs HSA. (C-D) For SEM pictures, the neutrophils cultured on a cover grass for 1 hour at 37°C under different conditions (HSA, HRG, AT, rAT, ARG, or HBSS) were fixed with 4% paraformaldehyde for 1 hour and then post fixed with 1% osmium overnight. The SEM pictures were obtained. Typical cell appearance is shown from each group. (E) The purified neutrophils cultured on a cover grass for 1 hour at 37°C under different conditions (HSA, HRG, AT, or HBSS) were fixed with 4% paraformaldehyde for 15 minutes. The neutrophils stained with phalloidin (F-actin; red), DNase I (G-actin; green), and Hoechst 33342 (nuclei; blue) were observed under a confocal laser scanning microscope. For panels A-B, original magnification, ×20; scale bar, 25 μm. For panels C-D, original magnification, ×12 000; scale bars, 5 μm. For panel E, original magnification, ×40; scale bar, low-magnification, 25 μm; high-magnification, 10 μm. For panels F-I, the results shown are the means ± SE of 12 fields.