Product Includes | Product # | Quantity | Color | Storage Temp |
---|---|---|---|---|
Inflammation Multi-Target | 97579 | 96 tests |
|
+4C |
TMB Substrate | 7004 | 11 ml |
|
+4C |
STOP Solution | 7002 | 11 ml |
|
+4C |
Sealing Tape | 54503 | 2 ea |
|
+4C |
ELISA Wash Buffer (20X) | 9801 | 25 ml |
|
+4C |
ELISA Sample Diluent | 11083 | 25 ml |
|
+4C |
Cell Lysis Buffer (10X) | 9803 | 15 ml |
|
-20C |
*The microwell plate is supplied as 12 8-well modules - Each module is designed to break apart for 8 tests.
Description
CST's PathScan® Inflammation Multi-Target Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that combines the reagents necessary to detect endogenous levels of NF-κB p65, phospho-NF-κB p65 (Ser536), phospho-SAPK/JNK (Thr183/Tyr185), phospho-p38 MAPK (Thr180/Tyr182), phospho-Stat3 (Tyr705) and phospho-IκB-α (Ser32). These molecules represent convergence points and key regulatory proteins in signaling pathways controlling the stress and inflammation response. Sixteen tests are provided for each target protein. Specific assay formulations for the indicated target proteins can be found in the datasheets associated with the individual PathScan® Sandwich ELISA Kits**. Briefly, a capture antibody* has been coated onto the microwells. After incubation with cell lysates, the coated antibody captures the target protein. Following extensive washing, a detection antibody* is added to detect the captured target protein. An HRP-linked secondary antibody is then used to recognize the bound detection antibody. HRP substrate, TMB, is added to develop color. The magnitude of absorbance for this developed color is proportional to the quantity of bound target protein. *Antibodies in kit are custom formulations specific to kit. **See companion products.
Specificity/Sensitivity
Background
Transcription factors of the nuclear factor κB (NF-κB)/Rel family play a pivotal role in inflammation, stress and immune responses. There are five family members in mammals: RelA/p65, c-Rel, RelB, NF-κB1 (p105/p50) and NF-κB2 (p100/p52). These proteins function as dimeric transcription factors. In unstimulated cells, NF-κB/Rel proteins are sequestered in the cytoplasm and inhibited by the IκB proteins. NF-κB-activating agents induce phosphorylation of IκB's, targeting them for degradation and thereby releasing the NF-κB/Rel complexes. Active NF-κB/Rel complexes are further activated by phosphorylation (1-4).
The stress-activated protein kinase/Jun-amino-terminal kinase SAPK/JNK is activated by a variety of environmental stresses, including UV and gamma radiation, ceramides, inflammatory cytokines and in some instances, by growth factors and GPCR agonists (5-10). As with the other MAPKs, the core-signaling unit is composed of a MAPKKK, typically MEKK1-4, or by a mixed lineage kinase (MLK), which phosphorylates and activates MKK4-7, which then phosphorylates Thr183 and Tyr185 to activate the SAPK/JNK kinase (6). Stress signals are delivered to this cascade by small GTPases of the Rho family (Rac, Rho, cdc42) (7). Both Rac1 and cdc42 mediate the stimulation of MEKKs and MLKs (7). Alternatively, MKK4-7 can be activated by a pathway independent of small GTPases via stimulation of a member of the germinal center kinase (GCK) family (8).
p38 MAP kinase (MAPK) participates in a signaling cascade controlling the cellular response to pro-inflammatory cytokines and a variety of cellular stresses. MKK3, MKK6 and SEK (MKK4) activate p38 MAP kinase by phosphorylation at Thr180 and Tyr182 (11-14).
The Stat3 transcription factor is an important signaling molecule for many cytokines and growth factor receptors. Stat3 is activated by phosphorylation at Tyr705, which induces dimerization, nuclear translocation and DNA binding (15,16).
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Background References
Cross-Reactivity Key
H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected
Trademarks and Patents
Limited Uses
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