Journal: Annals of Biomedical Engineering

Article Title: Contractility Dominates Adhesive Ligand Density in Regulating Cellular De-adhesion and Retraction Kinetics

doi: 10.1007/s10439-010-0195-z

Figure Lengend Snippet: Cytoskeletal architecture, focal adhesions, NMMII, and pMLC localization. U373 MG cells cultured overnight in serum-free media on FN-coated substrates of different densities were treated with 1 μ M LPA for 10 min and immunostained for F-actin, vinculin, NMMII, and pMLC, respectively. Under serum-free conditions, glioma cells exhibit a predominantly peripheral actin staining with few central stress fibers. Vinculin staining at focal adhesions and NMMII were diffused both in controls and in LPA-treated cells across all the different densities. pMLC was mostly diffusely distributed in untreated cells, but sometimes localized to peripheral membrane ruffles in LPA-treated cells (white arrows). Scale bar = 20 μ m

Article Snippet: Cells were incubated for 1 h at room temperature in one of the following primary antibody solutions in PBS at the following dilutions: mouse anti-vinculin (1:200, Sigma), rabbit anti-NMMII (1:200 anti-NMMIIA and 1:200 anti-NMMIIB, Sigma), and phosphorylated MLC (pMLC) (1:500, Cell Signaling).

Techniques: Cell Culture, Staining, Membrane