Lck (pY192) glyc Antibody | catalog# XBP-4157
Background
Lck (p56lck), a member of the Src family of non-receptor tyrosine protein kinases, is expressed predominantly in T-cells. Lck function is critical for both T-cell development in the thymus and activation of mature T-cells in the periphery by antigen. The activity of Lck is regulated by phosphorylation of multiple sites, including the two conserved residues, tyrosines 394 (equivalent to 418 in Src) and 505 (equivalent to 529 in Src). Phosphorylation of tyrosine 192 by Syk promotes activation of Lck by preventing the binding of the negative regulatory site, tyrosine 505.
Description
Background extracts alone (1) or with added recombinant active human Lck protein (2-5) were resolved by SDS-PAGE on a 10% Tris-glycine gel and transferred to PVDF. The membrane was blocked with a 5% BSA-TBST buffer for one hour at room temperature, and then incubated with the Lck [pS192] antibody in a 1% BSA-TBST buffer for two hours at room temperature, following prior incubation with: no peptide (1, 2), the non-phosphopeptide corresponding to the phosphopeptide immunogen (3), a generic phosphotyrosine-containing peptide (4) or the phosphopeptide immunogen (5). After washing, the membrane was incubated with goat F(ab')2 anti-rabbit IgG HRP conjugate and signals were detected using the Pierce SuperSignal™ method.
Specifications
Formulation
Rabbit polyclonal immunoglobulin in Dulbecco's phosphate buffered saline (without Mg2+ and Ca2+), pH 7.3 (+/- 0.1), 50% glycerol with 1.0 mg/mL BSA (IgG, protease free) as a carrier.Clonality/Clone
This is a polyclonal antibody.Specificity
Human Lck. Other species/Src family kinases that have not been tested include: mouse Src (100% homologous with the immunogen), chicken Src (79% homologous); Lck (79% homologous), Hck (79% homologous) and Lyn (93% homologous).Source
Lck antibody was produced against a chemically synthesized phosphopeptide derived from the region of human Lck that contains tyrosine 192 (based on Swiss Protein database, accession number P06239). The sequence is conserved in mouse.Purification
Lck antibody was purified from rabbit serum by sequential epitope-specific chromatography. The antibody has been negatively preadsorbed using i) a non-phosphopeptide corresponding to the site of phosphorylation to remove antibody that is reactive with non-phosphorylated Lck, ii) the phosphorylated peptide corresponding to Src tyrosine 215, and iii) a generic tyrosine phosphorylated peptide to remove antibody that is reactive with phosphotyrosine (irrespective of the sequence). The final product is generated by affinity chromatography using a Lck-derived peptide that is phosphorylated at tyrosine 192.Host
Lck (pY192) glyc antibody was raised in rabbit.Please use anti-rabbit secondary antibodies.
Application
For Western blotting applications, we recommend using the antibody at a 1:1000 starting dilution.Tested Application
WBBuffer
Rabbit polyclonal immunoglobulin in Dulbecco's phosphate buffered saline (without Mg2+ and Ca2+), pH 7.3 (+/- 0.1), 50% glycerol with 1.0 mg/mL BSA (IgG, protease free) as a carrier.Storage
Store at -20˚C. We recommend a brief centrifugation before opening to settle vial contents. Then, apportion into working aliquots and store at -20˚C. For shipment or short-term storage (up to one week), 2 - 8˚C is sufficient.Positive Control
- Full length untagged recombinant active human Lck protein.]
Species Reactivity
H, MReference
- Budagian, V., et al. (2003) Signaling through P2X7 receptor in human T cells involves p56(lck), kinases, and transcription factors AP-1 and NF-kappa B. J. Biol. Chem. 278(3):1549-1560.
- Mustelin, T. and K. Tasken (2003) Positive and negative regulation of T-cell activation through kinases and phosphatases. Biochem. J. 371(Pt 1):15-27.
- Goldmann, W.H. (2002) p56(lck) Controls phosphorylation of filamin (ABP-280) and regulates focal adhesion kinase (pp125(FAK)). Cell Biol. Int. 26(6):567-571.
- Gervais, F.G. and A. Veillette (1997) Reconstitution of interactions between protein-tyrosine phosphatase CD45 and tyrosine-protein kinase p56(lck) in nonlymphoid cells. J. Biol. Chem. 272(19):12754-12761.]