Phosphoproteomics - Overview of Phosphoproteomic Analysis

Overview of Phosphoproteomic Analysis

A sample large-scale phosphoproteomic analysis includes

  1. Cultured cells undergo SILAC encoding.
  2. Cells are stimulated with factor of interest (e.g. growth factor, hormone).
  3. Stimulation can occur for various lengths of time for temporal analysis.
  4. Cells are lysed and enzymatically digested.
  5. Peptides are separated using ion exchange chromatography.
  6. Phosphopeptides are enriched using phosphospecific antibodies, immobilized metal affinity chromatography or titanium dioxide (TiO2) chromatography.
  7. Phosphopeptides are analyzed using mass spectrometry.
  8. Peptides are sequenced and analyzed.

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