Time-lapse Fluorescence Microscopy of Neutrophil Extracellular Traps or NETs: A Technique to Analyze Post-adhesion Interaction between Cancer Cells and NETs

Culture and stain the peritoneal low-density neutrophils in a 35-millimeter dish, poly-L-lysine coated dish as demonstrated, followed by a brief wash in 2 milliliters of 0.1% BSA plus RPMI 1640.

Replace the medium with 1 times 10 to the sixth unstained gastric tumor cells suspended in 2 milliliters of RPMI supplemented with BSA for a 5-minute incubation at room temperature. At the end of the incubation, rinse the cells in 2 milliliters of fresh RPMI supplemented with BSA for 5 minutes at room temperature, followed by gentle swirling to remove any unattached tumor cells.

Then, replace the supernatant with DMEM supplemented with FBS and antibiotics and mount the dish in a whole-view cell observation system for time lapse analysis of tumor cell trapping by the NETs.