Product Pathways - PathScan ELISA
PathScan® Total Src Sandwich ELISA Kit #7984
| Kit Includes | Volume | Solution Color |
|---|---|---|
| Src Rabbit Antibody Coated Microwells | 96 tests | |
| Src Mouse Detection Antibody | 11 milliliters | Green |
| Anti-mouse IgG, HRP-linked Antibody | 11 milliliters | Red |
| Sample Diluent | 25 milliliters | Blue |
| TMB Substrate | 11 milliliters | Colorless |
| STOP Solution | 11 milliliters | Colorless |
| Sealing Tape | 2 sheets | |
| 20X Wash Buffer | 25 milliliters | Colorless |
| Cell Lysis Buffer (10X) # 9803 | 15 milliliters | 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 (10X), which is stored at –20°C (packaged separately).
Species Cross-Reactivity
H M
Reactivity Key: H=Human M=Mouse
Description
The PathScan® Total Src Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of total Src protein. A Src rabbit antibody has been coated onto the microwells. After incubation with cell lysates, Src protein (phospho and nonphospho) is captured by the coated antibody. Following extensive washing, a Src mouse detection antibody is added to detect the captured Src protein. Anti-mouse IgG, HRP-linked antibody is then used to recognize the bound detection antibody. The HRP substrate TMB is added to develop color. The magnitude of the absorbance for this developed color is proportional to the quantity of total Src protein.
Specificity / Sensitivity
The PathScan® Total Src Sandwich ELISA Kit #7984 from Cell Signaling Technology detects endogenous levels of total Src protein, as shown in Figure 1. This kit does not cross-react with the Src family members Yes, Lck, Lyn or Hck. Other family members have not been tested. The kit sensitivity is shown in Figure 2.
ELISA
Figure 1. Constitutive phosphorylation of Src at Tyr416 in Calu-3 cells as detected by the PathScan® Phospho-Src (Tyr416) Sandwich ELISA Kit #7953. Treatment of lysate with λ-phosphatase significantly reduces Src phosphorylation but does not alter the levels of total Src protein as detected by the PathScan® Total Src Sandwich ELISA kit #7984. Absorbance at 450 nm is shown in the top figure while the corresponding western blots using Src (L4A1) Mouse mAb #2110 (left) or Phospho-Src Family (Tyr416) (100F9) Rabbit mAb #2113 (right) are shown in the bottom figure.
ELISA
Figure 2. The relationship between lysate protein concentration from untreated and λ-phosphatase treated lysate from Calu-3 cells and the absorbance at 450 nm is shown.
Background
The Src family of protein tyrosine kinases, which includes Src, Lyn, Fyn, Yes, Lck, Blk and Hck, are important in the regulation of growth and differentiation of eukaryotic cells (1). Src activity is regulated by tyrosine phosphorylation at two sites, but with opposing effects. Phosphorylation of Src at Tyr416 in the activation loop of the kinase domain by carboxy-terminal Src kinase (Csk) upregulates enzyme activity, whereas phosphorylation of Tyr527 in the carboxy-terminal tail renders the enzyme less active (2).
- Thomas, S.M. and Brugge, J.S. (1997) Annu. Rev. Cell Dev. Biol. 13, 513-609.
- Hunter, T. (1987) Cell 49, 1-4.
Application References
Have you published research involving the use of our products? If so we'd love to hear about it. Please let us know!
Companion Products
- 2109 Src (36D10) Rabbit mAb
- 2123 Src (32G6) Rabbit mAb
- 9803 Cell Lysis Buffer (10X)
- 7076 Anti-mouse IgG, HRP-linked Antibody
- 7004 TMB Substrate
- 7002 STOP Solution
- 9808 Phosphate Buffered Saline (PBS-20X)
- 9809 Phosphate Buffered Saline with Tween 20 (PBST-20X)
This product is intended for research purposes only. The product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
