TCA PRECIPITATION OF PROTEINS

(Protocol prepared by Lori Kohlstaedt, June 2006) of the Mass Spectrometry Facility of the University of California at Berkeley, it is reproduced here for your conveneince..

Reagents

Preparation of 100% TCA: (don’t try to weigh out TCA; it’s too hygroscopic)

• Obtain a fresh bottle of crystalline TCA.
• Read the weight in the container from the label.
• Add distilled water to give a 100 g / 100 ml solution at final volume.

0.01 M HCl/90% acetone

Procedure:

1. Add 100% (w/v) TCA (trichloroacetic acid, ( see preparation method above) to the sample to bring the TCA concentration to 20%.
2. Incubate on ice for at least 1 hr. Diluted samples may be left overnight.
3. Spin at maximum speed in a microcentrifuge for 10 min.
4. Wash the pellet 3X with a solution of ice cold 0.01 M HCl / 90% acetone. Allow the pellet to air dry.
   The pellet can then be resuspended directly in 100 mM Tris, pH 8.5, 8 M urea or 6 M Guanidine- HCl in 25 mM ammonium bicarbonate buffer for enzymatic digestion for mass spectrometry