*5.11. Cell Harvest and Western Blot Analysis*

Cells were washed twice with ice-cold PBS and lysed with cell lysis buffer containing 5 µL/mL protease inhibitor cocktail in IGEPAL. Lysates were syringed with 23-gauge needles to shear cellular DNA and centrifuged at 12,000× *g* rpm at 4 ◦C for 5 min. Total cell protein concentrations of the resulting supernatants were determined using BCA protein assays. Equal amounts of cell protein were loaded onto 10% (*v*/*v*) acrylamide gels for separation by SDS-PAGE. Proteins were transferred onto nitrocellulose membranes, blocked for 1 h using 5% (*w*/*v*) skim milk or 0.1% (*w*/*v*) BSA in PBS-Tween (0.05% *v*/*v*), then incubated with anti-ABCB1 (1:2000 (Novus Biologicals), overnight 4 ◦C) or antitubulin (1:3000, overnight 4 ◦C) antibodies. Membranes were rinsed with PBS-Tween, then incubated with HRP-conjugated anti-mouse secondary antibodies (1:10,000) for 1 h at room temperature, and then rinsed again with PBS-Tween. Protein bands were visualised via chemiluminescence and the Bio-Rad ChemiDoc imaging system.
