Rabbit anti actin

HT-29 cells were grown on coverslips for 24 h and treated with 20 nM dasatinib for 24 h. The cells were fixed with 3.7 % formaldehyde for 20 min and permeabilized with 0.1 % Triton X-100 for 1 min. The fixed cells were incubated with mouse anti-E-cadherin (BD Biosciences), rabbit anti-p-Src (Cell Signaling Technology), rabbit anti-actin (GeneTex), rabbit anti-Rab11 (Invitrogen), rabbit anti-pEGFR (Invitrogen) and rabbit anti-p-FAK (Cell Signaling Technology) for 1 h at room temperature, followed by incubation with secondary antibodies conjugated to Alexa Fluor 488 or 594 (Jackson ImmunoResearch) for 1 h at room temperature. After washing with PBS, the coverslips were mounted with Fluoromount (Merck-Sigma, MA, USA), and images were acquired using a Zeiss LSM 510 META confocal system with a 63X objective (1.4 oil). Z-scanning sections were taken for 3D imaging, and line scans and side views were analyzed using Zeiss LSM image software. For quantification of E-cadherin, Rab11 and pEGFR distribution at leading edge and the cell-cell contacts, more than 100 cells of each independent experiment were counted, the positive counts were divided by the total cell-cell contact numbers.