Cell Signaling Technology

Product Pathways - PathScan ELISA

PathScan® Phospho-Stat3 (Ser727) Chemiluminescent Sandwich ELISA Kit #8026

When ordering five or more kits, please contact us for processing time and pricing at sales@cellsignal.com.

Kit Includes Volume Solution Color
Stat3 Rabbit mAb Coated Microwells 96 tests
P-Stat3 (S727) Mouse Detection Antibody 5.5 ml Green
Anti-mouse IgG, HRP-linked Antibody 5.5 ml Red
Luminol/Enhancer Solution 3 ml Colorless
Stable Peroxide Buffer 3 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 Cross-Reactivity

H

Reactivity Key:  H=Human
Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Description

The PathScan® Phospho-Stat3 (Ser727) Chemiluminescent Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of phospho-Stat3 (Ser727) protein with a chemiluminescent readout. Chemiluminescent ELISAs often have a wider dynamic range and higher sensitivity than conventional chromogenic detection. This chemiluminescent ELISA, which is offered in low volume microplates, shows increased signal and sensitivity while using smaller samples. A Stat3 Rabbit mAb has been coated on the microwells. After incubation with cell lysates, Stat3 (phospho or nonphospho) protein is captured by the coated antibody. Following extensive washing, a Phospho-Stat3 (Ser727) Mouse mAb is added to detect the captured phospho-Stat3 protein. Anti-mouse IgG, HRP-linked Antibody is then used to recognize the bound detection antibody. Chemiluminescent reagent is added for signal development. The magnitude of light emission, measured in relative light units (RLU), is proportional to the quantity of phospho-Stat3 (Ser727) protein.Antibodies in kit are custom formulations specific to kit.

Specificity / Sensitivity

PathScan® Phospho-Stat3 (Ser727) Chemiluminescent Sandwich ELISA Kit detects endogenous levels of phospho-Stat3 (Ser727) in human cells. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

Sensitivity

Sensitivity

Figure 1. Relationship between protein concentration of lysates from untreated and hEGF-treated A-431 cells and immediate light generation with chemiluminescent substrate is shown. Cells (85% confluence) were treated with hEGF #8916 (100 ng/ml) and lysed after incubation at 37ºC for 5 minutes. Graph inset corresponding to the shaded area shows high sensitivity and a linear response at the low protein concentration range.

Background

The Stat3 transcription factor is an important signaling molecule for many cytokines and growth factor receptors (1) and is required for murine fetal development (2). Research studies have shown that Stat3 is constitutively activated in a number of human tumors (3,4) and possesses oncogenic potential (5) and anti-apoptotic activities (3). Stat3 is activated by phosphorylation at Tyr705, which induces dimerization, nuclear translocation, and DNA binding (6,7). Transcriptional activation seems to be regulated by phosphorylation at Ser727 through the MAPK or mTOR pathways (8,9). Stat3 isoform expression appears to reflect biological function as the relative expression levels of Stat3α (86 kDa) and Stat3β (79 kDa) depend on cell type, ligand exposure, or cell maturation stage (10). It is notable that Stat3β lacks the serine phosphorylation site within the carboxy-terminal transcriptional activation domain (8).

  1. Heim, M.H. (2001) J Recept Signal Transduct Res 19, 75-120.
  2. Takeda, K. et al. (1997) Proc Natl Acad Sci U S A 94, 3801-4.
  3. Catlett-Falcone, R. et al. (1999) Immunity 10, 105-15.
  4. Garcia, R. and Jove, R. (1998) J Biomed Sci 5, 79-85.
  5. Bromberg, J.F. et al. (1999) Cell 98, 295-303.
  6. Darnell, J.E. et al. (1994) Science 264, 1415-21.
  7. Ihle, J.N. (1995) Nature 377, 591-4.
  8. Wen, Z. et al. (1995) Cell 82, 241-50.
  9. Yokogami, K. et al. (2000) Curr Biol 10, 47-50.
  10. Biethahn, S. et al. (1999) Exp Hematol 27, 885-94.

Application References

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Protocols

* Product-specific protocol.

Companion Products

For Research Use Only. Not For Use In Diagnostic Procedures.

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