Welcome to Gaia! :: View User's Journal | Gaia Journals

Migration assay. Wound healing assays were used to evaluate the migration of 95D Walrycin B transfected with siRNA against CXCR4 (termed as 95D-siR-CXCR4 cells) and 95D cells transfected with control siRNA (termed as 95D-ctrl-CXCR4 cells). Cells were seeded in six-well plates at 1 × 105 per well in growth medium. Confluent monolayers were starved overnight in assay medium and a single scratch wound was created using a micropipette tip. The cells were washed with PBS to remove cell debris, supplemented with assay medium containing 10 μg/ml CpG ODN and monitored. Images were captured with a microscope using a 5× objective at and 24 h post-wounding. For a quantitative measure, we counted the number of migrating cells to wound area based on the picture at h post-wounding.
Animal experiments. 95D cells (5 × 106) transfected with CXCR4–siRNA (treated group) or control-siRNA (control group) were pretreated with 10 μg/ml CpG ODN. After 3 days, the cell were collected and then injected into each mouse of 16 nude mice (8 mice each group) by the tail vein. At 30 days post-injection of cells, the mice were sacrificed. Lungs were harvested in optimum cutting temperature compound and frozen in liquid nitrogen. The collected lung tissue sections were subjected to H&E histostaining. The metastasis foci also were counted under microscope.