Product namePhospho-JNK (T183/Y185) + Total In-Cell ELISA Kit (Chemiluminescent)
Sample typeAdherent cells, Suspension cells
Assay typeCell-based (quantitative)
Assay durationMultiple steps standard assay
Species reactivityReacts with: Mouse, Rat, Human
Phospho-JNK (T183/Y185) + Total In-Cell ELISA Kit (Chemiluminescent) (ab207477) provides a simple, efficient, cell-based method to monitor proteins activated by phosphorylation. The kit is designed specifically to quantify activated (phosphorylated) JNK and/or total JNK. Cells are cultured in 96-well plates and stimulated to induce the pathway of interest. Following stimulation, the cells are rapidly fixed to preserve activation-specific protein modifications. Each well is then incubated with a primary antibody that recognizes either phosphorylated JNK or total JNK. Subsequent incubation with secondary HRP-conjugated antibody and chemiluminescent reagent provides an easily quantified chemiluminescent readout. The relative number of cells in each well is then determined using the provided Crystal Violet solution. The 96-well plate format is suitable for high-throughput screening applications.
The Phospho-JNK (T183/Y185) + Total In-Cell ELISA Kit (Chemiluminescent) (ab207477) contains 96-well plates and two primary antibodies. The phospho-JNK antibody was raised against a synthetic phospho-peptide corresponding to the sequence surrounding Thr183 and Tyr185 of human JNK. This antibody recognizes JNK1 only when phosphorylated at these sites, and JNK2 and JNK3 only when phosphorylated at equivalent sites. The total-JNK antibody recognizes JNK1, JNK2 and JNK3 regardless of the phosphorylation state.
The kit can be used to study phosphorylated JNK relative to cell number or to determine JNK phosphorylation relative to the total JNK protein found in the cells. Once the phospho-JNK and total-JNK signals have been normalized for cell number, a comparison of the ratio of phosphorylated JNK to total JNK for each of the cell growth conditions can be made.
JNK (c-Jun N-terminal kinase), also called stress activated protein kinase (SAPK), is involved in a mitogen-activated protein (MAP) kinase cascade that transduces cellular stress signals like inflammation and apoptosis. JNK is activated by exposure of cells to environmental stress such as UV light and by the treatment of cells with cytokines. Activation of JNK is mediated by dual phosphorylation at Thr183 and Tyr185. This phosphorylation is performed by the MAP kinase kinases, which serve as signaling molecules that integrate a wide array of stimuli into the activation of the JNK signaling pathway.
Activated JNK phosphorylates and activates several transcription factors including c-Jun, ATF-2, Elk-1 and JunD. The activated JNK pathway in turn regulates AP-1 transcriptional activity in vivo and is required for embryonic morphogenesis, the regulation of cellular proliferation and apoptosis, and the response of cells to immunological stimuli.
In neurons, JNK regulates differentiation as well as apoptotic programs. Activation of the JNK pathway may contribute to the neuronal atrophy and death that is associated with neuro-degenerative pathological conditions including cerebral ischemia, Alzheimer’s, Parkinson’s and Huntington’s Diseases. In studies of tumor cells, JNK has been implicated in signaling cell survival and has been shown to play an important role in the pathogenesis of many proliferative diseases, including cancer.
The JNK pathway is also involved in T-lymphocyte activation and differentiation and mediates feedback inhibition of the insulin signaling cascade. Novel findings indicate that regulation of JNK activity is important in the control of osteoarthritis.
Elucidation of the signaling cascades mediated by JNK will provide insight into the delicate balance between cellular life and death and aid in drug discovery efforts.
Tested applicationsSuitable for: In-Cell ELISAmore details
Storage instructionsPlease refer to protocols.
Components 1 x 96 tests 5 x 96 tests 1% SDS Solution 1 x 22ml 5 x 22ml 10% Triton X-100 1 x 10ml 5 x 10ml 10X PBS 1 x 120ml 5 x 120ml 1X Antibody Blocking Buffer 1 x 22ml 5 x 22ml 1X Antibody Dilution Buffer 1 x 30ml 5 x 30ml 96-well tissue culture plate 2 units 10 units Anti-rabbit HRP-conjugated IgG 1 x 11µl 5 x 11µl Chemiluminescent Reagent 2 x 2ml 10 x 2ml Crystal Violet Solution 1 x 22ml 5 x 22ml Phospho-JNK antibody 1 x 9µl 5 x 9µl Plate sealer 2 units 10 units Reaction Buffer 2 x 4ml 10 x 4ml Total-JNK antibody 1 x 9µl 5 x 9µl
Cellular localizationJNK1+JNK2+JNK3: Cytoplasm. Nucleus.
Our Abpromise guarantee covers the use of ab207477 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|In-Cell ELISA||Use at an assay dependent concentration.|
This product has been referenced in:
- Liu J et al. CD9 regulates keratinocyte migration by negatively modulating the sheddase activity of ADAM17. Int J Biol Sci 15:493-506 (2019). Read more (PubMed: 30745837) »
- Chen X et al. Evodiamine alleviates severe pneumonia induced by methicillin-susceptible Staphylococcus aureus following cytomegalovirus reactivation through suppressing NF-?B and MAPKs. Int J Mol Med 42:3247-3255 (2018). Read more (PubMed: 30320369) »