Isolation of neutrophils

The Institutional Review Board of the Sixth Affiliated Hospital of Guangzhou Medical University approved this work. Neutrophils were isolated from heparinized blood of healthy donors aged 25–35 years by using the Human Peripheral Blood Neutrophil Isolation Kit (Beyotime, Shanghai, China) according to the manufacturer’s instructions. During isolation of the cells, pyrogen contamination and excessive mechanical stimulation were carefully avoided. The average purity of neutrophils was > 95%, as determined by Wright-Giemsa staining, and the average viability of isolated cells was > 98%, as assessed by trypan blue dye exclusion. After isolation, the cells were resuspended in Ca²⁺-free HBSS at 5 × 10⁶ cells/ml and stored at room temperature until use (< 1 h).