Immunofluorescent analysis of Phospho-MAPK1/3 pThr202/Tyr204 (green) in HeLa cells either left untreated (left panel) or treated with 50nM PMA (right panel) for 10 minutes. Formalin fixed cells were permeabilized with 0.1% Triton X-100 in TBS for 10 minutes at room temperature and blocked with 1% Blocker BSA (Product # 37525) for 15 minutes at room temperature. Cells were probed with a Phospho-MAPK1/3 pThr202/Tyr204 polyclonal antibody (Product # PA1-4607) at a dilution of 1:100 for at least 1 hour at room temperature, washed with PBS, and incubated with DyLight 488 goat anti-rabbit IgG secondary antibody (Product # 35552) at a dilution of 1:400 for 30 minutes at room temperature. F-Actin (red) was stained with DyLight 554 Phalloidin (Product # 21834) and nuclei (blue) were stained with Hoechst 33342 dye (Product # 62249). Images were taken on a Thermo Scientific ArrayScan or ToxInsight Instrument at 20X magnification.
|Tested species reactivity||Human, Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic phospho-peptide corresponding to amino acid residues surrounding Thr202/Tyr204 of rat MAPK1/2 conjugated to KLH|
|Storage buffer||0.01M HEPES, pH 7.5, with 0.15M NaCl, 100µg/ml BSA, 50% glycerol|
|Storage Conditions||-20° C, Avoid Freeze/Thaw Cycles|
|Tested Applications||Dilution *|
|Immunohistochemistry (Paraffin) (IHC (P))||Assay-Dependent|
|Western Blot (WB)||1:1,000|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
This antibody is predicted to react with bovine, canine, chicken, mouse, non-human primate, Xenopus and zebrafish based on 100% sequence homology.
In Western blot, this antibody detects an ~42-44 kDa protein representing Phospho-Thr202/Thr204 ERK/MAPK.
Antibodies to this protein (and modification) were previously sold as part of a Thermo Scientific Cellomics High Content Screening Kit. Further optimization, including the selection of the most appropriate fluorescent Dylight conjugated secondary antibody, may have to be performed for your high content assay.
Extracellular-Signal Regulated Kinase/Mitogen-Activated Protein Kinase (ERK/MAPK) is an integral component of cellular signaling during mitogenesis and differentiation of mitotic cells and also is thought to play a key role in learning and memory. The activity of this kinase is regulated by dual phosphorylation at Thr202 and Tyr204.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.