Revision 12

#9101

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Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Applications:
W, W-S, IP, IF-IC, FC-FP

Reactivity:
H M R Hm Mk Mi Dm Z B Pg Ce

Sensitivity:
Endogenous

MW (kDa):
42, 44

Source/Isotype:
Rabbit

UniProt ID:
#P27361, #P28482

Entrez-Gene Id:
5595, 5594

Product Usage Information

Application Dilution
Western Blotting 1:1000
Simple Western™ 1:10 - 1:50
Immunoprecipitation 1:50
Immunofluorescence (Immunocytochemistry) 1:200 - 1:800
Flow Cytometry (Fixed/Permeabilized) 1:100 - 1:400

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.

Specificity/Sensitivity

Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody detects endogenous levels of p44 and p42 MAP Kinase (Erk1 and Erk2) when phosphorylated either individually or dually at Thr202 and Tyr204 of Erk1 (Thr185 and Tyr187 of Erk2). This antibody does not cross-react with the corresponding phosphorylated residues of either JNK/SAPK or p38 MAP Kinase, and does not cross-react with non-phosphorylated Erk1/2.

Species predicted to react based on 100% sequence homology

Chicken

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Thr202/Tyr204 of human p44 MAP kinase.

Background

Mitogen-activated protein kinases (MAPKs) are a widely conserved family of serine/threonine protein kinases involved in many cellular programs, such as cell proliferation, differentiation, motility, and death. The p44/42 MAPK (Erk1/2) signaling pathway can be activated in response to a diverse range of extracellular stimuli, including mitogens, growth factors, and cytokines (1-3), and research investigators consider it an important target in the diagnosis and treatment of cancer (4). Upon stimulation, a sequential three-part protein kinase cascade is initiated, consisting of a MAP kinase kinase kinase (MAPKKK or MAP3K), a MAP kinase kinase (MAPKK or MAP2K), and a MAP kinase (MAPK). Multiple p44/42 MAP3Ks have been identified, including members of the Raf family, as well as Mos and Tpl2/COT. MEK1 and MEK2 are the primary MAPKKs in this pathway (5,6). MEK1 and MEK2 activate p44 and p42 through phosphorylation of activation loop residues Thr202/Tyr204 and Thr185/Tyr187, respectively. Several downstream targets of p44/42 have been identified, including p90RSK (7) and the transcription factor Elk-1 (8,9). p44/42 are negatively regulated by a family of dual-specificity (Thr/Tyr) MAPK phosphatases, known as DUSPs or MKPs (10), along with MEK inhibitors, such as U0126 and PD98059.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

W: Western Blotting W-S: Simple Western™ IP: Immunoprecipitation IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized)

Cross-Reactivity Key

H: Human M: Mouse R: Rat Hm: Hamster Mk: Monkey Mi: Mink Dm: D. melanogaster Z: Zebrafish B: Bovine Pg: Pig Ce: C. elegans

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

Alexa Fluor is a registered trademark of Life Technologies Corporation.

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 12

Cell Signaling Technology Logo
Western blot analysis of extracts from various cell lines treated with TPA (200nM, 4 hr) or PDGF (100ng/mL, 10 min) as indicated, using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody (upper), p44/42 MAPK (Erk1/2) (137F5) Rabbit mAb #4695 (middle), or β-Actin (D6A8) Rabbit mAb #8457 (lower).
Western Blotting Image 1: Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody
Specificity and sensitivity of Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody. The antibody reacts specifically with as little as 0.25 ng of phosphorylated p42 MAP kinase and does not cross-react with up to 4 µg of nonphosphorylated p42 MAP kinase.
Western Blotting Image 2: Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody
Western blot analysis of whole-cell extracts from unstarved wild-type mouse embryonic fibroblasts (MEFs) treated with the indicated combinations of basic Fibroblast Growth Factor (bFGF, 100 ng/ml for 30 minutes), Platelet-Derived Growth Factor (PDGF, 100 ng/ml for 30 minutes), MEK1 Inhibitor (PD98059 #9900, 50 µM, 2 hour pre-treatment), and MEK1/2 Inhibitor (U0126 #9903, 10 µM, 2 hour pre-treatment), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody #9101 (upper panel) and p44/42 MAPK (Erk1/2) (137F5) Rabbit mAb #4695 (lower panel).
Western Blotting Image 3: Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 12

Cell Signaling Technology Logo
Simple WesternTM analysis of lysates (0.1 mg/mL) from 3T3+PDGF cells using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) #9101.  The virtual lane view (left) shows the target band (as indicated) at 1:10 and 1:50 dilutions of primary antibody.  The corresponding electropherogram view (right) plots chemiluminescence by molecular weight along the capillary at 1:10 (blue line) and 1:50 (green line) dilutions of primary antibody.  This experiment was performed under reducing conditions on the JessTM Simple Western instrument from ProteinSimple, a BioTechne brand, using the 12-230 kDa separation module.
Western Blotting Image 1: Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody
Immunoprecipitation of Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) from Hela cell extracts. Cells were treated with TPA, (400 nM, 4 hrs). Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody. Western blot was performed using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody. Mouse Anti-rabbit IgG (Light-Chain Specific) (D4W3E) mAb #45262 was used as a secondary antibody.
Immunoprecipitation Image 1: Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody
Confocal immunofluorescent analysis of NIH/3T3 cells, treated with U0126 (10 uM, 2 hr; left) or treated with TPA (200 nM, 30 min; right), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody (green), β-Actin (8H10D10) Mouse mAb #3700 (red), and DAPI #4083 (blue).
Immunofluorescence Image 1: Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 12

Cell Signaling Technology Logo
Flow cytometric analysis of Jurkat cells, treated with U0126 #9903 (10 uM, 2 hr; blue) or treated with TPA (12-O-Tetradecanoylphorbol-13-Acetate) #4174 (200 nM, 30 min; green) using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')₂ Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Flow Cytometry Image 1: Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) Antibody
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.