Rabbit anti c jun

Cultured cells were collected and solubilized using protein lysis buffer. The proteins were then separated by size using SDS-PAGE and transferred to polyvinyl difluoride membranes (Millipore). The membranes were incubated with primary antibodies followed by secondary antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA). The immunoreactive proteins were detected using an enhanced chemiluminescence kit (Millipore). The primary antibodies were rabbit anti- E-cadherin, rabbit anti-N-cadherin, rabbit anti-Vimentin (Cell Signaling Technology, Danvers, MA, USA), mouse anti-MMP9, mouse anti-MMP2, mouse anti-JNK1, mouse anti-p-JNK, rabbit anti-c-Jun, mouse anti-p-c-Jun, mouse anti-FAK, mouse anti-β-actin (Santa Cruz Biotechnology), rabbit anti-Paxillin, and rabbit anti-p-Paxillin (Abcam, Cambridge, MA, USA). The digital images of the Western blotting bands were quantified by Meta Morph software (MDS Analytical Technologies) after the background subtraction.

Mouse cortical neurons at 7 days in primary culture were harvested with lysis buffer (10 mM Tris-HCl, 10 mM NaCl, 0.5 mM EDTA, 1% NP-40, and 0.5% protease inhibitor cocktail) and lysates were rotated for 60 min at 4 °C. After 20 min of centrifugation at 12,000 rpm, supernatants were incubated with 1 μg antibody for 12 h at 4 °C. Antibodies were used as follows; rabbit anti-SRm300 (Santa Cruz Biotechnology, sc-292291); mouse anti-PQBP1 (Santa Cruz Biotechnology, sc-374260); mouse-anti-SC35 (Sigma-Aldrich, S4045); rabbit anti-c-Jun (Santa Cruz Biotechnology, sc-44). Protein G Sepharose (GE Healthcare, Buckinghamshire, UK) was incubated for 4 h at 4 °C, and beads were washed with lysis buffer four times. Then, sample buffer was added to the beads, and samples were boiled at 95 °C for 10 min before SDS-PAGE.