Cell Signaling Technology

Product Pathways - PathScan ELISA

PathScan® Total SAPK/JNK Chemiluminescent Sandwich ELISA Kit #7869

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

Important Ordering Details: Product is assembled upon order to ensure maximum activity. Domestic: Please allow up to two weeks for your order to be processed and shipped. International: Please allow up to three weeks, depending on the country, for your order to be processed and shipped.

Kit Includes Volume Solution Color
SAPK/JNK Mouse mAb Coated Microwells 96 tests
SAPK/JNK Rabbit Detection mAb 5.5 ml Green
Anti-rabbit 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
Sample Diluent 25 ml Blue
Cell Lysis Buffer (10X) #9803 15 ml

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 M

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

Description

The PathScan® Total SAPK/JNK Chemiluminescent Sandwich ELISA Kit is a solid phase sandwich enzyme-linked immunosorbent assay (ELISA) that detects endogenous levels of total SAPK/JNK 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 a smaller sample size. A SAPK/JNK mouse mAb has been coated on the microwells. After incubation with cell lysates, SAPK/JNK proteins are captured by the coated antibody. Following extensive washing, a SAPK/JNK rabbit mAb is added to detect the captured SAPK/JNK protein. Anti-rabbit 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 total SAPK/JNK protein.Antibodies in kit are custom formulations specific to kit.

Specificity / Sensitivity

PathScan® Total SAPK/JNK Chemiluminescent Sandwich ELISA Kit #7869 detects endogenous levels of total SAPK/JNK in human and mouse cells. This kit detects proteins from the indicated species, as determined through in-house testing, but may also detect homologous proteins from other species.

Relationship between protein concentration of lysates from untreated and UV-treated 293 cells and immediate light generation with chemiluminescent substrate is shown. Cells (70-90% confluent) were treated with or without UV and lysed after incubation at 37ºC for 30 minutes. Graph inset corresponding to the shaded area shows high sensitivity and a linear response at the low protein concentration range.

Background

The stress-activated protein kinase/Jun-amino-terminal kinase SAPK/JNK is potently and preferentially activated by a variety of environmental stresses including UV and gamma radiation, ceramides, inflammatory cytokines, and in some instances, growth factors and GPCR agonists (1-6). As with the other MAPKs, the core signaling unit is composed of a MAPKKK, typically MEKK1-MEKK4, or by one of the mixed lineage kinases (MLKs), which phosphorylate and activate MKK4/7. Upon activation, MKKs phosphorylate and activate the SAPK/JNK kinase (2). Stress signals are delivered to this cascade by small GTPases of the Rho family (Rac, Rho, cdc42) (3). Both Rac1 and cdc42 mediate the stimulation of MEKKs and MLKs (3). Alternatively, MKK4/7 can be activated in a GTPase-independent mechanism via stimulation of a germinal center kinase (GCK) family member (4). There are three SAPK/JNK genes each of which undergoes alternative splicing, resulting in numerous isoforms (3). SAPK/JNK, when active as a dimer, can translocate to the nucleus and regulate transcription through its effects on c-Jun, ATF-2, and other transcription factors (3,5).

  1. Davis, R.J. (1999) Biochem Soc Symp 64, 1-12.
  2. Ichijo, H. (1999) Oncogene 18, 6087-93.
  3. Kyriakis, J.M. and Avruch, J. (2001) Physiol Rev 81, 807-69.
  4. Kyriakis, J.M. (1999) J Biol Chem 274, 5259-62.
  5. Leppä, S. and Bohmann, D. (1999) Oncogene 18, 6158-62.
  6. Whitmarsh, A.J. and Davis, R.J. (1998) Trends Biochem Sci 23, 481-5.

Application References

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Protocols

* Product-specific protocol.

Companion Products

Rabbit Monoclonals are produced under license (granting certain rights, including those under U.S. Patent No. 5,675,063) from Epitomics, Inc.

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

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