TCA/Acetone Precipitation

The following procedure is used to precipitate protein from freshly prepared cell lysates. This allows for optimal protein recovery and accurate assays. The acetone used in this procedure should be prechilled and stored in a -20°C freezer until needed and kept on ice during the entire procedure.

Procedure

1. Mix the following solutions in a 1:8:1 ratio in the following order (invert tube after adding each component):
                        1 ml cell lysate
                        8 ml 100% ice-cold acetone
                        1 ml 100% trichloroacetic acid (100% TCA)
2. Precipitate at -20° C for 1 hour. 
3. Centrifuge at 11,500 rpm (18,000 x g) for 15 min at 4°C in a microcentrifuge.
4. Discard supernatant. Wash with 1 ml ice-cold acetone. Centrifuge at 11,500 rpm for 15 min at 4°C.
5. Discard supernatant.
6. Air dry  the pellet and go for solution digest.

(Sudhakar Voruganti protocol Jan. 2014)