Figure 4

Comparison of the 3D invasion assay and the Boyden chamber invasion assay. A. MDA-MB-231, LM2-4 and HT1080 cells infected with lentiviruses encoding the control shRNA SHC002 (sh-control) or lentiviruses encoding the Loxl2 specific shRNA TRCN46195 (sh-Loxl2) were seeded as monolayers in collagen as described under Figure 1. After 4 days the gels frozen and were sectioned perpendicular to the cell monolayer. Shown are cryostat sections in which the cells were stained with eosin-Y. Three independent experiments were performed with similar results. B. The number of tumor cells infected with a control shRNA or with the Loxl2 specific shRNA encoding lentiviruses was determined as described in methods. Each histogram represents the average number of invading cells determined from examination of eight to twelve microscopic fields derived from three different experiments. C. Representative pictures of HT1080 fibrosarcoma cells infected with lentiviruses encoding control (sh-control) or Loxl2 targeting (sh-Loxl2) shRNA species that have invaded and crossed the filter of Boyden chambers is shown. The histogram shows the average number of cells that transversed the Boyden chamber filter in two independent experiments. Error bars represent the standard error of the mean. *** p < 0.001.