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TCA precipitation for SILAC-labeled cells

Author: Tao Tao Chen, updated date: , view: 6, Q&A: 0
Tags: SILAC and TCA

Procedure

TCA precipitation



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

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

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

  4. Incubate in ice 10 min

  5. Centrifuge at 4 C at max speed 10 min

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

  7. Vortex 2X ~10 sec

  8. Incubate at -20 C 5min

  9. Spin 15 max speed at 4 C

  10. Wash again with 500ul cold acetone

  11. Vortex lightly

  12. Spin 15 min at 4 C

  13. Remove acetone

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

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