plasma protein precipitation

[varshaparekh]

Hi all,
Does anybody have a protocol for precipitation of plasma proteins from human blood

[victor]

1. Use RBC lysis Buffer about 3x blood volume and incubate for 10 minutes and centrifuge at 1500 rpm; 10 minutes (rt) and after that, pour the supernatant --> do this 3 times.
2. Vortex the pellet and use Cell lysis solution (about 625 uL for 2,5 mL blood) & 2,5 uL RNase --> incubate for 30 minutes; 37C.
3. Do the protein precipitation by adding 5M ammonium acetate (about 417 uL for 2,5 mL blood sample) and vortex it until the solution looks white milky.
4. Centrifuge at 3000 rpm, 4C, 15 minutes and pour the supernatant.
5. There you'll get blood protein and plasma protein.

Hope this helps.
Victor