2.5 Transwell migration/invasion assay
The migration and invasion abilities of pancreatic cancer cells were
determined by using Transwell Boyden Chambers with 8-μm pores (Corning
Inc., Lowell, MA, USA). The upper Transwell chambers were uncoated for
cell migration assay or pre-coated with 50 µL of Matrigel (1:4 mixed
with PBS; BD Biosciences, San Jose, CA, USA) for cell invasion assay.
The cells were precultured in serum-free medium for 12 h. Next, the
cells were harvested and resuspended in serum-free medium containing
serial concentrations of TA, and then plated on the upper chambers at an
appropriate density. At the same time, complete medium containing 10%
FBS was added to the lower chambers. After an incubation of 48 h, the
tumor cells on the upper side of the filters were wiped off with cotton
swabs. The tumor cells on the lower surface of the membrane were then
fixed with 4% paraformaldehyde and stained with 0.1% crystal violet.
The images of stained tumor cells were acquired using an inverted
microscope (Olympus, Tokyo, Japan).