The PTMScan^® Control Peptides Phospho-Tyrosine enable quality control of immunoaffinity enrichment performance using PTMScan^® or PTMScan^® HS workflows.  These synthetic peptides contain a specific post-translational modification (PTM) that can be enriched by the associated PTMScan^® or PTMScan^® HS immunoaffinity purification (IAP) beads, as well as a stable heavy isotope that can be distinguished from endogenous peptides by the mass spectrometer.

Tyrosine phosphorylation plays a key role in cellular signaling (1). Research studies have shown that in cancer, unregulated tyrosine kinase activity can drive malignancy and tumor formation by generating inappropriate proliferation and survival signals (2). Antibodies specific for phospho-tyrosine (3,4) have been invaluable reagents in these studies. The phospho-tyrosine monoclonal antibodies developed by Cell Signaling Technology are exceptionally sensitive tools for studying tyrosine phosphorylation and monitoring tyrosine kinase activity in high throughput drug discovery.

Use with Cell Signaling Technology’s PTMScan^® kit protocol from the Immunoaffinity Purification (IAP) step. Because the optimal amount of PTMScan^® Control Peptides Phospho-Tyrosine for each user’s experiments will depend on unique factors, such as mass spectrometer sensitivity, users may dilute these control peptides as needed.

1. Aliquot PTMScan^® Control Peptides Phospho-Tyrosine for storage as single-use units at -20°C or proceed to immediate usage.
2. Resuspend sample peptides in the appropriate buffer and volume, e.g., 1.4 mL of PTMScan^® IAP Buffer (1X).
3. Clear sample peptides by centrifugation.
4. Transfer clarified sample peptides to tubes containing IAP beads.
5. Add 10 µL of PTMScan^® Control Peptides Phospho-Tyrosine to IAP beads and sample peptides and mix well.
6. Continue with PTMScan^® or PTMScan^® HS workflows at the 2-hour incubation step.
7. Detect PTMScan^® Control Peptides Phospho-Tyrosine in the LCMS data file.