Analysis Note

ControlEGF-stimulated A431 cell lysate.

Included Positive Antigen Control:old> Catalog # 12-302, EGF-stimulated A431 cell lysate is provided as a free positive antigen control for western immunoblotting. Aliquot as desired, refreeze immediately, and store at -20°C. The lysate is stable for 6 months at -20°C. Before use, add 2.5 µL of 2-mercaptoethanol/100 µL of lysate and boil for 5 minutes to reduce the preparation. old>Load 20 µg of reduced lysate per lane for immunoblot analysis.

Application

Anti-Phosphotyrosine Antibody, clone PY20 is an antibody against Phosphotyrosine for use in ELISA, IP & WB.

Immunoprecipitation:
Recommended by an independent laboratory to immunoprecipitate tyrosine phosphorylated proteins.

ELISA:
Recommended by an independent laboratory.

General description

Tyrosine phosphorylation represents a major mechanism for the activation of signal transduction pathways in response to growth factors, hormones and cytokines. There are approximately 100 tyrosine kinases that are both receptor tyrsosine kinase receptors for various growth factors and cytokines. The extent and duration of tyrosine phosphorylation are tightly regulated by the 107 protein tyrosine phosphatases. Tyrosine phosphorylation occurs on residues on either other kinases (Ser/Thr or Tyr) or on substrates. In the case of kinases, this phosphorylation often causes the enzyme to change activation states and/or it creates binding sites for SH2 domain. Mutation or misregulation of protein tyrosine kinases is a cause or consequence of many disease states. It has been estimated that 20% of oncogenes so far described are tyrosine kinases. With such clear links between aberrant protein tyrosine kinase activity and disease, it is no surprise that much of the early effort in kinase drug discovery was focused on these enzymes.

Immunogen

Phosphotyrosine conjugated to carrier protein.

Legal Information

UPSTATE is a registered trademark of Merck KGaA, Darmstadt, Germany

Linkage

Replaces: MAB3080

Other Notes

Concentration: Please refer to the Certificate of Analysis for the lot-specific concentration.

Physical form

Format: Purified

0.1M Tris-Glycine, 0.15M NaCl, 0.05% Sodium Azide, pH 7.4. Liquid at 2-8°C.

Quality

Routinely evaluated by western blot with RIPA lysates EGF-treated human A431 carcinoma cells.

Western Blot Analysis:
0.5-2 µg/mL of this lot detected tyrosine-phosphorylated proteins in RIPA lysates EGFtreated human A431 carcinoma cells.

Specificity

Reactivity with other species has not been confirmed.

Storage and Stability

Stable for 1 year at 2-8°C from date of recipt.

NOTE: DO NOT FREEZE.old>
For maximum recovery of the product, centrifuge the original vial prior to removing the cap. If the product has accidentally been frozen and thawed, spin it at 13,000 x g for 10 minutes at 2-8°C. Save the supernatant for application.

Target description

Dependent upon the molecular weight of the tyrosine phosphorylated protein being detected.