Western Blotting Protocol
2. Transfer protein onto a Shleicher & Schuell Nitrocellulose membrane (0.4 u) 1 X Transfer Buffer with a BioRad Semi-Dry Transferring Cell 1 h at 12 V.
3. Block in 1X TBST containing 5% milk for 1 h at RT or ON at 4oC with agitation. .
4. Add anti-serum to a proper amount of 1 X TBST containing 5% milk (enough to cover the membrane) at 1:2000 or 1:1000 dilution. Shake the blot 1 to 2 h at RT.
5. Wash the blot with 1X TBST 3 times. Every time 5 minutes at RT.
6. Incubate the blot with PIERCE Immuno Pure Rabbit Anti-mouse IgG + IgM (H + L), HRP Conjugated, at a 1:5000 dilution for 1 h at RT.
7. Wash 3 times as step 5.
8. Incubate the blot with SuperSignal West Pico Chemiluminescent Substrate (Product # 34080) as described by the manufacturer.