Immunoblotting analysis of mechano-sensitive molecular constituents of the focal adhesion complex in PDL cells after strain-application. PDL cells were seeded on flexible bottom cell culture dishes, strained with averaged 2.5% and followed by westernblot analysis. The numerical expression values were always denoted in relation to unstrained controls. (A) Expression levels of integrin β3 in PDL cells in response of the strain kinetic were detected with a mouse monoclonal anti-integrin β3 antibody and the maximum expression modulation was numerical notified in the graph after 6 h of strain. (B) For detection of the strain-induced total protein expression level of the focal adhesion kinase (FAK), a polyclonal rabbit anti-FAK antibody was used, and the maximum expression value was marked in the graph after 6 h strain. (C) Phosphorylation levels of Tyr576 of FAK were assessed by immunoblotting using an antibody against phospho-Tyr576. The maximum activation status was numerically denoted after 0.25 h strain. (D) In addition, for loading control and for internal normalisation, β-actin was detected for each immunoblotting experiment. Data of each graph represent the mean of three individual experiments (n = 3), mean +/- SD and means were subjected to the Students T-test. All compared mean values with p < 0.01 were considered as statistical significant and are marked with an asterisk.
Ziegler et al. BMC Cell Biology 2010 11:10 doi:10.1186/1471-2121-11-10