TCA precipitation for SILAC-labeled cells

Make 100% TCA, 3g in 3mls of filtered water, ice it

Mix SILAC samples: L, M, H together in equal volumes or equal protein amount

Add TCA to a final concentration of 10%, i.e.: 60ul of TCA in 600ul sample

Incubate in ice 10 min

Centrifuge at 4 C at max speed 10 min

Wash with 500 ul cold acetone (store acetone at -20 C)

Vortex 2X ~10 sec

Incubate at -20 C 5min

Spin 15 max speed at 4 C

Wash again with 500ul cold acetone

Vortex lightly

Spin 15 min at 4 C

Remove acetone

Evaporate in hood 1-2 min (don’t dry pellet completely)