Boyden cell migration assay

Tumor lysates (1 mg/mL) or mouse recombinant mouse CXCL2 (50 ng/mL; R&D; 452-M2) were prepared in HBSS and used in neuroprobe cell migration chamber with 3-µm filter. Mice were treated with vehicle control or either CXCR2 antagonist for more than 2 d prior to inducing sterile peritonitis by injecting 1 mL of 4% thioglycolate intraperitoneally. Peritoneal neutrophils were collected from peritoneal exudates 4 h after the induction of sterile peritonitis. Hemocytometer counts and modified Wright stained cytospins were utilized to confirm >85% neutrophil purity. The cells were used in the migration chamber in HBSS at 2 × 10⁶ cells/mL. After 3 h, the migrated cells were counted by flow cytometry.