Wesrern Blot Protocol

1. Load ~50 micrograms of lysate (ie: liver membrane) onto an 8% SDS-PAGE, reducing.

2. Block the protein transferred membrane for 30 minutes in blocking buffer [5% NFDM, 10 mM Tris (pH 7.6) saline, 0.2% Tween-20] at room temperature.

3. Incubate the membrane in diluted primary anti-ABCG8 [cat# NB 400-110], 1:500 for 1 hour at room temperature.

4. Wash the membrane 3x 5 minutes in blocking buffer without milk.

5. Incubate the membrane in diluted secondary anti-Rabbit IgG conjugated to HRP.

6. Wash the membrane 3x 5 minutes in blocking buffer without milk.

7. Wash the membrane for 5 minutes in TBS.

8. Develop using West-Pico ECL (Pierce) reagent.