Figure 6

Biological validation of expression datasets at the protein level. (A) Confocal analysis of HMLE-SNAIL and Kras-HMLE-SNAIL cells post GN-25 treatment. Cell were exposed to indicated concentrations of GN-25 for 24 hrs in 4 well chambered slides and immunofluorescence assay was performed according to published methods . Slides were stained with either vimentin (cell signaling), Snail (Cell signaling) or E-Cadherin (cell signaling) antibodies overnight. Secondary Antibody staining was performed for 2 hrs using anti-mouse Alexafluor antibody (Invitrogen). (B) Western blot analysis of HMLE-SNAIL and Kras-HMLE-SNAIL cells exposed to indicated concentrations of GN-25 for 24 hrs. The blots were probed for TWIST1, TWIST2 and E-Cadherin. GAPDH was used as a loading control. Blots are representative of two independent experiments.