Fig. 6. Paxillin expression is maintained following differentiation of NMuMG cells in a collagen I gel. NMuMG cells were cultured under normal 2D conditions or cultured in a 3D collagen I gel for a period of 7 days. (A) Hoffman modulation contrast images (20x) were taken of NMuMG cells cultured under 2D conditions (a) and under 3D collagen gel conditions (b). (B) Detergent-soluble lysates (DSL) were prepared from 2D and 3D gel-cultured NMuMG cells and were processed for western blot analysis. Appropriate antibodies were utilized to detect the indicated proteins and their phosphorylated isoforms. (C) Paxillin was immunoprecipitated from 2D and 3D gel lysates and immunoblotted for paxillin and phospho-Y118 paxillin. Although paxillin expression levels remain unchanged, basal paxillin phospho-Y118 was absent from cells cultured under 3D collagen gel conditions. Bar, 10 µm.