| 货号 | 7345C |
| 描述 | CSTs PathScan® Phospho-PDGF Receptor beta (Tyr751) Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of Phospho-PDGF Receptor beta (Tyr751) protein. A PDGF Receptor beta (7345-28D12) Rabbit mAb* has been coated onto the microwells. After incubation with cell lysates, Both nonphospho- and phospho- PDGF Receptor beta proteins are captured by the coated antibody. Following extensive washing, Phospho-PDGF Receptor beta Mouse mAb #3166* is added to detect the captured phospho-PDGF Receptor beta protein. HRP-linked anti-mouse antibody #7076* is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of optical density for this developed color is proportional to the quantity of phospho-PDGF Receptor beta (Tyr751) protein. * Antibodies in the kit are custom formulations specific to the kit |
| 反应种属 | Human/Mouse |
| 应用 | ELISA |
| 目标/特异性 | CSTs PathScan® Phospho-PDGF Receptor beta (Tyr751) Sandwich ELISA Kit detects endogenous levels of Phospho-PDGF Receptor beta (Tyr751) protein. Using this Sandwich ELISA Kit #7345, a significant induction of phospho-PDGF Receptor beta (Tyr751) in NIH/3T3 cells treated with PDGF can be detected. However, the level of total PDGF Receptor beta protein remains unchanged, as shown by Western analysis using PDGF Receptor beta Rabbit mAb (7345-28D12) [Fig.1]. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species. |
| 供应商 | CST |
| 背景 | The proteins of the platelet derived growth factor (PDGF) family exist as several disulphide-bonded, dimeric isoforms (PDGF AA, PDGF AB, PDGF BB, PDGF CC, and PDGF DD) that bind in a specific pattern to two closely related receptor tyrosine kinases, PDGF receptor α (PDGFRα) and PDGF receptor β (PDGFRβ). PDGFRα and PDGFRβ share 75% to 85% sequence homology between their two intracellular kinase domains while the kinase insert and carboxy-terminal tail regions display a lower level (27% to 28%) of homology (1). PDGFRα homodimers bind all PDGF isoforms except those containing PDGF D. PDGFRβ homodimers bind PDGF BB and DD isoforms, as well as the PDGF AB heterodimer. The heteromeric PDGF receptor α/β binds PDGF B, C, and D homodimers as well as the PDGF AB heterodimer (2). PDGFRα and PDGFRβ can each form heterodimers with EGFR, which is also activated by PDGF (3). Various cells differ in the total number of receptors present and in the receptor subunit composition, which may account for responsive differences among cell types to PDGF binding (4). Ligand binding induces receptor dimerization and autophosphorylation, followed by binding and activation of cytoplasmic SH2 domain-containing signal transduction molecules such as Grb2, Src, GAP, PI3 kinase, PLCγ, and Nck. A number of different signaling pathways are initiated by activated PDGF receptors and lead to control of cell growth, actin reorganization, migration, and differentiation (5). Tyr751 in the kinase-insert region of PDGFRβ is the docking site for PI3 kinase (6). Phosphorylated pentapeptides derived from Tyr751 of PDGFRβ (pTyr751-Val-Pro-Met-Leu) inhibit the association of the carboxy-terminal SH2 domain of the p85 subunit of PI3 kinase with PDGFRβ (7). Tyr740 is also required for PDGFRβ-mediated PI3 kinase activation (8). |
| 存放说明 | 4C |
| 参考文献 | Deuel, T.F. et al. (1988) Biofactors 1, 213-217. Bergsten, E. et al. (2001) Nat. Cell Biol. 3, 512-516. Betsholtz, C. et al. (2001) Bioessays 23, 494-507. Coughlin, S.R. et al. (1988) Prog. Clin. Biol. Res. 266, 39-45. Ostman, A. and Heldin, C.H. (2001) Adv. Cancer Res. 80, 1-38. Panayotou, G. et al. (1992) EMBO J. 11, 4261-4272. Ramalingam, K. et al. (1995) Bioorg. Med. Chem. 3, 1263-1272. Kashishian, A. et al. (1992) EMBO J. 11, 1373-1382. |
Figure 1: Treatment of NIH/3T3 cells with PDGF stimulates phosphorylation of PDGF Receptor beta at Tyr751, detected by PathScan® Phospho-PDGF Receptor beta (Tyr751) Sandwich ELISA kit, #7345, but does not affect the level of total PDGF Receptor beta protein detected by PDGF Receptor beta Rabbit mAb (7345-28D12) via Western analysis. OD450 readings are shown in the top figure, while the corresponding Western blot using Phospho-PDGF Receptor beta (Tyr751) (88H8) Mouse mAb #3166 (right panel) or PDGF Receptor beta Rabbit mAb (7345-28D12) (left panel), is shown in the bottom figure.图1：用PathScan® Phospho-PDGF Receptor beta (Tyr751) Sandwich ELISA kit, #7345能够在PDGF刺激后的NIH/3T3细胞中检测到PDGF Receptor beta上Tyr751位点的磷酸化，但不影响用PDGF Receptor beta Rabbit mAb (7345-28D12)对总PDGF Receptor beta的免疫印迹检测结果。上图显示OD450nm下的读值，下图显示用Phospho-PDGF Receptor beta (Tyr751) (88H8) Mouse mAb #3166（右道）或PDGF Receptor beta Rabbit mAb (7345-28D12)（左道）进行免疫印迹检测的结果。 | |
Figure 2: Linear relationship between protein concentration of lysates from control or PDGF-treated NIH/3T3 cells and kit assay optical density readings. NIH/3T3 cells (75% confluence) were treated with PDGF (50 ng/ml), and lysed after incubation at 37°C for 5 minutes.图2：NIH/3T3细胞经过和未经PDGF刺激后其裂解液中蛋白含量和试剂盒检测读值之间的线性关系。用PDGF (50 ng/ml)在37°C 刺激NIH/3T3细胞（铺至75%满）5分钟，然后裂解细胞。 |
