Transwell chamber dishes

Transwell chamber dishes (8 μm, Corning) were used to perform the tumor invasion assay. Raji cells (1 × 10⁵ cells/well) were placed in the upper chamber, while immature macrophages (6 × 10⁴ cells/well) were placed in the lower chamber. After incubation for 48 h, non-migratory cells on the upper surface of the insert were gently removed using a cotton-tipped swab, and cells that entered the lower surface of the membrane were fixed and stained with crystal violet at room temperature for 20 min. Light microscopy was used to count cells that invaded through the Matrigel coating. For quantification, five randomly selected fields on the lower side of the insert were photographed. Each experiment was performed in triplicate.

Co-culture experiments were performed, as previously reported [17 (link), 18 (link)]. Raji cells (5 × 10⁴ cells/well) and immature macrophages (6 × 10⁴ cells/well) were co-cultured in DMEM medium (Gibco) supplemented with 2% FBS for 72 h in 0.4-μm Transwell chamber dishes (Corning). For amphotericin B treatment, immature macrophages were first treated with 1 μM of amphotericin B for 72 h, and subjected to co-culture experiments without amphotericin B. In order to rule out the effect of amphotericin B directly on Raji cells, cells washed once after drug treatment.