This antibody specifically recognizes β-catenin phosphorylated at tyrosine 654 at 90 kDa and gamma-catenin at 70 kDa. Due to sequence homologies of the phospho-epitope surrounding pY 654 there may be crossreactions with other proteins (e.g. Annexin A11 at 56 kDa, gamma-catenin at 70 kDa, Desmoplakin at 81 kDa, TNFalpha-induced protein 2 at 73 kDa, Ribosomal protein S6 kinase alpha 6 at 90 kDa).
Western Blot: approx.1 μg/mL for HRPO/ECL detection. Recommended buffer: Casein/Tween 20 based blocking and blot incubation buffer. Positive control: Cell lysate from pervanadate-treated SW480 cells (please see: 'Protocols') Other applications not tested. Optimal dilutions are dependent on conditions and should be determined by the user.
Protocol
Positive Control Cell Lysate: SW 480 PERVANADATE TREATEDFormat: Lyophilized cell lysate from SW480 cellsSerum starved cells were treated for 15 min with pervanadate. Reconstitution: Restore by addition of 200 µl H2O. After complete solubilization add 200 µl 2x SDS-PAGEsample buffer, mix and incubate at 90°C for 5 min. Application: The positive control cell lysate is recommended for immunoblot applications. 20 µl ofpositive control cell lysate correspond to ca. 20. 000 cells. Use 20 µl/lane (mini gel) for HRPO/ECL detection of the target proteins. Please NOTE: The lyophilized cell lysates conatin SDS and are not recommended forapplications with native proteins such as in immunoprecipitation. Storage: Aliquote reconstituted product and store frozen. Avoid repeated fereezing and thawing.
Restrictions
For Research Use only
Reconstitution
Restore with 1 mL H2O (15 min, RT).
Buffer
2 x PBS containing 0.09 % Na-azide, PEG and Sucrose.
Preservative
Sodium azide
Precaution of Use
WARNING: Reagents contain sodium azide. Sodium azide is very toxic if ingested or inhaled. Avoid contact with skin, eyes, or clothing. Wear eye or face protection when handling. If skin or eye contact occurs, wash with copious amounts of water. If ingested or inhaled, contact a physician immediately. Sodium azide yields toxic hydrazoic acid under acidic conditions. Dilute azide-containing compounds in running water before discarding to avoid accumulation of potentially explosive deposits in lead or copper plumbing.
Storage
-20 °C
Storage Comment
For long-term storage, freeze lyophilizate upon arrival (-20 °C). Upon reconstitution,aliquote and freeze in liquid nitrogen, reconstituted antibody should be stored frozen at-80 °C. Avoid repeated freeze-thaw cycles. Shelf life: one year from despatch. Thaw aliquots at 37 °C. Thawed aliquots may be stored at 4 °C up to 3 months.
The α-, β- and γ-catenins are cytoplasmic proteins mediating the interaction of Ca2+-dependent transmembrane adhesion molecules (cadherins) with the cytoskeletal network. Additionally, β-catenin is a substrate of both receptor and non-receptor tyrosine kinases. Tyrosine 86 and tyrosine 654 are substrates of EGF receptor and src family kinases while tyrosine 142 is a substrate of fer tyrosine kinases.Synonyms: Beta-catenin, CTNNB, CTNNB1