What protocol should I use to prepare serum lysates for western blot?

First, ensure blood is collected in a vacutainer tube that does not contain any anticoagulants such as sodium heparin or EDTA.
1. In a biosafety hood transfer the blood into an appropriately sized conical tube or 1.5mL Eppendorf tube. Let the blood sit at room temperature for a least 1 hour until the blood clots.
2. Spin the tube at 1400rpm for 5 minutes. The resulting supernatant is the serum. Since the blood has had time to clot, the serum should be free of blood cells, fibrinogen, and platelets. Be careful not to disturb the pellet when pipetting off the serum.
3. Mix one part serum with one part 2X SDS/DTT. Sonicate the lysate and then boil for 5 minutes.

Last updated: September 12, 2024