| 货号 | 14541T |
| 目标/特异性 | Unless otherwise indicated, each antibody will recognize endogenous total levels of target protein, and modification state antibodies will only recognize target proteins phosphorylated at the indicated residue. Phospho-Lck (Tyr505) Antibody may cross-react with certain phosphorylated Src family members due to high sequence homology. Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb may cross-react with other Src family members (Lyn, Fyn, Lck, Yes and Hck) when phosphorylated at equivalent sites, and may cross react with overexpressed phosphorylated RTKs. Phospho-Zap-70 (Tyr319)/Syk (Tyr352) (65E4) Rabbit mAb cross-reacts with endogenous levels of Syk when phosphorylated at Tyr352. Phospho-Zap-70 (Tyr493)/Syk (Tyr526) Antibody cross-reacts with endogenous levels of Syk when phosphorylated at Tyr526. |
| 供应商 | CST |
| 背景 | When T cells encounter antigens via the T cell receptor (TCR), information about the quantity and quality of antigens is relayed to the intracellular signal transduction machinery (1). This activation process depends mainly on CD3 (Cluster of Differentiation 3), a multiunit protein complex that directly associates with the TCR α and ß chains. CD3 is composed of four polypeptides: ζ, γ, ε and δ. Each of these polypeptides contains at least one immunoreceptor tyrosine-based activation motif (ITAM) (2). The Src family kinases Lck and Fyn are recruited to the TCR complex upon stimulation and activate the downstream tyrosine kinases to initiate signaling. Phosphorylation of Lck at Tyr394 leads to an increase in Lck activity while phosphorylation of Tyr505 in the Lck carboxy-terminal tail down-regulates Lck catalytic activity (3). Zap-70 and Syk are rapidly phosphorylated on several tyrosine residues through autophosphorylation and transphosphorylation by Src family tyrosine kinases. Activation loop phosphorylation of Zap-70 at Tyr493 and Syk at Tyr526 leads to complete activation of both kinases (4). Subsequent phosphorylation of other tyrosine residues within the kinase interdomain B region, including Zap-70 at Tyr315 and Zap-70 at Tyr 319, create docking sites for downstream signaling molecules. Zap-70 and Syk phosphorylate the transmembrane adaptor protein LAT at multiple, conserved tyrosine residues within SH2 binding motifs, exposing these motifs as docking sites for downstream signaling targets (5,6). The phosphorylation of LAT at Tyr171 and Tyr191 enables the binding of Grb2, Gads/SLP-76, PLCγ1, and PI3 kinase. The adapter protein SLP-76 is phosphorylated at Tyr113 and Tyr128, allowing for binding of the Grb2-like adapter Gads. Phosphorylation of SLP-76 at Ser376 by hematopoietic progenitor kinase 1 (HPK1) induces interaction with 14-3-3ε and down-regulates TCR signaling (7,8). Phosphoinositide-specific phospholipase PLCγ1 enzyme activity is also stimulated by Zap-70 and Syk phosphorylation on Tyr783, Tyr711, and Tyr1253, resulting in robust PI-4,5-P2 hydrolysis (9). |
| 存放说明 | -20C |
| 参考文献 | Kuhns, M.S. et al. (2006) Immunity 24, 133-9. Pitcher, L.A. and van Oers, N.S. (2003) Trends Immunol 24, 554-60. Chow, L.M. et al. (1993) Nature 365, 156-60. Wang, H. et al. (2010) Cold Spring Harb Perspect Biol 2, a002279. Zhang, W. et al. (1998) Cell 92, 83-92. Paz, P.E. et al. (2001) Biochem J 356, 461-71. Shui, J.W. et al. (2007) Nat Immunol 8, 84-91. Di Bartolo, V. et al. (2007) J Exp Med 204, 681-91. Beach, D. et al. (2007) J Biol Chem 282, 2937-46. |
Western blot analysis of extracts from Jurkat cells (starved for 16 hours) treated with calf intestinal alkaline phosphatase (CIP) or H2O2 (2 mM), using Phospho-Lck (Tyr505) Antibody (upper) or control Lck Antibody #2752 (lower). | |
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition. | |
Western blot analysis of extracts from Jurkat cells starved for 16 hours and treated with 2 mM H2O2 for 3 minutes, using Phospho-Zap-70 (Tyr493)/Syk (Tyr526) Antibody (upper) or control Zap-70 Antibody #2702 (lower). | |
Western blot analysis of SDS extracts from untreated or anti-CD3 treated (10 µg/ml for 2 minutes) human Jurkat cells after overnight serum starvation using Phospho-LAT (Tyr191) Antibody. | |
Western blot analysis of extracts from Jurkat cells treated with hydrogen peroxide (2mM for 2 minutes) or with lambda phosphatase and extracts from Ramos cells treated with anti-human IgM (12 micrograms/ml for 2 minutes) using Phospho-Zap-70 (Tyr319)/Syk (Tyr352) (65E4) Rabbit mAb. | |
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGlo® is added and emits light during enzyme catalyzed decomposition. | |
Western blot analysis of total cell lysates from DND41 and Molt4 cells using CD3ε (CD3-12) Rat mAb. | |
Flow cytometric analysis of Jurkat cells using CD3ε (CD3-12) Rat mAb Antibody (blue) compared to a nonspecific negative control antibody (red). | |
Western blot analysis of extracts NIH/3T3 cells, serum-starved or treated with human Platelet-Derived Growth Factor BB hPDGF-BB #8912 (100 ng/ml, 15 min), using Phospho-Src Family (Tyr416) (D49G4) Rabbit mAb (upper) or Src (36D10) Rabbit mAb #2109 (lower). | |
Western blot analysis of extracts from Ramos cells untreated or treated with 10 µg/ml IgM for 2 minutes with or without calf intestinal phosphatase (CIP), using Phospho-Zap-70 (Tyr493)/Syk (Tyr526) Antibody (upper) or Syk Antibody #12358 (lower). | |
Western blot analysis of extracts from Jurkat cells, untreated (-) or treated with H2O2 (11 mM, 1 min; +), using Phospho-SLP-76 (Ser376) Antibody (upper) or SLP-76 Antibody #4958 (lower). | |
Western blot analysis of extracts from EL4 cells, untreated (-) or treated with H2O2 (11 mM, 1 min; +), using Phospho-SLP-76 (Ser376) Antibody (upper) or SLP-76 Antibody #4958 (lower). | |
Flow cytometric analysis of NIH/3T3 cells, untreated (blue) or treated with recombinant mouse PDGF-BB (200 ng/ml, 15 min; green), using Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb. Anti-rabbit IgG (H+L), F(ab)2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody. | |
Confocal immunofluorescent analysis of A-431 cells, untreated (left), treated with hEGF #8916 (100 ng/ml, 5 min; center), or treated with hEGF #8916 (100 ng/ml, 5 min) and λ phosphatase (right), using Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye). | |
Immunoprecipitation of phospho-PLCγ1 (Tyr783) from A-431 cell extracts stimulated with hEGF #8916 using Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (lane 2) or Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb (lane 3). Lane 1 is 10% input. Western blot analysis was performed using Phospho-PLCγ1 (Tyr783) (D6M9S) Rabbit mAb. |
