Western Blot Protocol:
1. SDS electrophoresis
2. Transfer (use Immobilon PVDF or Hybond nitro-cellulose (Amersham)
  - activate in methanol for 1-2 minutes
  - wash with water for 5 minutes
  - incubate in transfer buffer for 5 minutes (15,1g TRIS, 72,1g glycine, 1000ml methanol, add water up to final 5000ml, pH8,5)
  - bring gel/membrane/3MM paper together
  - transfer over night (330 mA for 28x18 cm gels) or equivalent
3. Immunoblot
  - block with PBS/10% non-fat dry milk/0,3% Tween 20 over night at 4°C or 1 hour at 37°C
  - wash 2x with PBS/0,3% Tween 20 for 5 minutes
  - 10µg/ml in PBS/5% non-fat dry milk/0,3% Tween 20
-> If you do not see anything at this concentration there is not enough detectable protein, under normal conditions 1µg/ml may be sufficient.
  - incubate 1 hour at room temperature
  - wash 4x with PBS/0,3% Tween 20 for 5 minutes
- second antibody: rabbit anti-mouse-HRPO (DAKO) 1:1’000 in PBS/5% non-fat dry milk/0,3% Tween 20 for 1hour at room temperature
  - wash 4x with PBS/0,3% Tween 20 for 5 minutes
4. detection with ECL system from Amersham
  - mix sol1 and sol2 in equal amounts
  - incubate blot for 1 minute at room temperature in this mixture
  - dry blot carefully with tissue
  - cover blot in Saran Wrap
  - lay "side of gel" to X-ray film, expose for 5 seconds to 10 minutes.