*4.11. Western Blot Analysis*

The cells were pre-seeded in 6 well culture plates at 2 × 10<sup>5</sup> cells mL−<sup>1</sup> for 24 h and treated with di fferent sample concentrations. After 24 h, the cells were harvested, lysed, and centrifuged at 12,000× *g* for 20 min to remove the cellular debris. The proteins were resolved by SDS-polyacrylamide jell electrophoresis. Protein bands were blotted onto nitrocellulose membranes and incubated for 2 h with skim milk in TBST. The membranes were consequently incubated with primary (8 h under 4 ◦C) and secondary antibodies (2 h at room temperature). Protein bands were visualized by adding chemiluminescent substrate (Cyanagen Srl, Italy) using a FUSION SOLO Vilber Lourmat system, France [21].
