Product Pathways - PathScan ELISA
PathScan® Phospho-PDGF Receptor β (Tyr751) Sandwich ELISA Kit #7345
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|Kit Includes||Volume||Solution Color|
|PDGF Receptor beta Rabbit mAb Coated Microwells|
|P-PDGF Receptor beta (Y751) Detection Ab||11 ml||Green|
|Anti-mouse IgG, HRP-linked Antibody||11 ml||Red|
|TMB Substrate #7004||11 ml||Colorless|
|STOP Solution #7002||11 ml||Colorless|
|Sealing Tape||2 sheets|
|ELISA Wash Buffer (20X)||25 ml||Colorless|
|ELISA Sample Diluent||25 ml||Blue|
|Cell Lysis Buffer (10X) #9803||15 ml||Yellowish|
Note: 12 8-well modules –Each module is designed to break apart for 8 tests.
Note: Kit should be stored at 4°C with the exception of Cell Lysis Buffer, which is stored at –20°C (packaged separately).
Species enclosed in parentheses are predicted to react based on 100% sequence homology.
CST's 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 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 is added to detect the captured phospho-PDGF Receptor beta protein. Anti-mouse IgG, HRP-linked 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 kit are custom formulations specific to kit.
Specificity / Sensitivity
CST's 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.
ELISA - Western correlation
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.
Platelet derived growth factor (PDGF) family proteins 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).
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- Bergsten, E. et al. (2001) Nat. Cell Biol. 3, 512-516.
- Betsholtz, C. et al. (2001) Bioessays 23, 494-507.
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- 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.
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- 3164 PDGF Receptor α Antibody
- 3162 PDGF Receptor β Antibody
- 3166 Phospho-PDGF Receptor β (Tyr751) (88H8) Mouse mAb
- 3161 Phospho-PDGF Receptor β (Tyr751) Antibody
Rabbit Monoclonals Produced UsingTechnology from Epitomics, Inc. UnderU.S. patent No. 5,675,063.
For Research Use Only. Not For Use In Diagnostic Procedures.