Quick diff stain kit

The cell invasion assay was carried out according to our previous study [66 (link)] using 10-well chemotaxis chambers (Neuro Probe, Gaithersburg, Maryland, USA) with an 8-µM pore membrane (Neuro Probe) in RPMI-1640 with 10% FBS as the chemoattractant in the lower chamber. Briefly, AGS cells were added to the upper chamber with nicotine for 24 h, the non-invading cells on the upper surface of each membrane were removed from the chamber by using cotton swabs, and the invading cells on the lower surface of each membrane were stained using the Quick-Diff stain kit (Becton-Dickinson, Franklin Lakes, NJ, USA). After two washes with water, the chambers were allowed to air dry. The number of invading cells was counted using a phase-contrast microscope.

The cell invasion assay was carried out using the 10-well chemotasis chamber (Neuro Probe, Gaithersburg, Maryland, USA) with 8 μM pore membrane (Neuro Probe) with 10% FBS containing RPMI as the chemoattractant in the lower chamber. AGS cells (10⁵ in 280 μl) were added to upper chamber with PMA in the presence of chrysin, MMP-9 antibody or nonspecific IgG, and incubated to invade the Matrigel for 24 hours. In order to determine the effect of chrysin and signaling inhibitors on PMA-induced cell invasion, AGS cells were pre-incubated with chrysin, curcumin, PD98059, or SP600125 for 1 hour, and incubated with PMA for the 24 hour invasion period. The non-invading cells on the upper surface of each membrane were removed from the chamber, and the invading cells on the lower surface of each membrane were stained with the Quick-Diff stain kit (Becton-Dickinson, Franklin Lakes, NJ, USA). After two water washes, the chambers were allowed to air-dry. The number of invading cells was counted using a phase-contrast microscope.

The cell invasion assay was carried out using 10-well chemotaxis chambers (Neuro Probe, Gaithersburg, Maryland, USA) with an 8-μM pore membrane (Neuro Probe) in DMEM with 10% FBS as the chemoattractant in the lower chamber. The non-invading cells on the upper surface of each membrane were removed from the chamber by using cotton swabs, and the invading cells on the lower surface of each membrane were stained using the Quick-Diff stain kit (Becton-Dickinson, Franklin Lakes, NJ, USA). After two washes with water, the chambers were allowed to air dry. The number of invading cells was counted using a phase-contrast microscope.