Product Pathways - MAPK Signaling

Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb #4370

This XP™ monoclonal antibody was developed using our eXceptional Monoclonal Technology (XMT™).

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Applications	Reactivity	Sensitivity	MW (kDa)	Isotype
W IP IHC-P IF-IC F	H M R Hm Mk Mi Dm Z B Dg Pg Sc (Ce)	Endogenous	44, 42	Rabbit IgG

Applications Key: W=Western Blotting IP=Immunoprecipitation IHC-P=Immunohistochemistry (Paraffin) IF-IC=Immunofluorescence (Immunocytochemistry) F=Flow Cytometry
Reactivity Key: H=Human M=Mouse R=Rat Hm=Hamster Mk=Monkey Mi=Mink Dm=D. melanogaster Z=Zebrafish B=Bovine Dg=Dog Pg=Pig Sc=S. cerevisiae Ce=C. elegans
Species cross-reactivity is determined by Western blot.

Protocols

4370:: Flow, IHC / Paraffin, Immunofluorescence, Immunoprecipitation, Western Blotting

Specificity / Sensitivity

Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb detects endogenous levels of p44 and p42 MAP Kinase (Erk1 and Erk2) when dually phosphorylated at Thr202 and Tyr204 of Erk1 (Thr185 and Tyr187 of Erk2), and singly phosphorylated at Thr202. This antibody does not cross-react with the corresponding phosphorylated residues of either JNK/SAPK or p38 MAP kinases.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Thr202/Tyr204 of human p44 MAP kinase.

Western Blotting

Western blot analysis of extracts from COS cells, untreated or treated with either U0126 #9903 (10 µM for 1h) or TPA #9905 (200 nM for 10 m), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb #4370 and p44/42 MAPK (Erk1/2) (3A7) Mouse mAb #9107.

Western Blotting

Western blot analysis of extracts from 293, NIH/3T3 and C6 cells, treated with λ phosphatase or TPA as indicated, using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb (upper), or p44/42 MAPK (Erk1/2) (137F5) Rabbit mAb #4695 (lower).

IHC-P (paraffin)

Immunohistochemical analysis using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb on SignalSlide™ Phospho-p44/42 MAPK (Thr202/Tyr204) IHC Controls #8103 (paraffin-embedded NIH/3T3 cells, untreated (left) or PMA-treated (right).

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb.

IHC-P (paraffin)

Immunohistochemical analysis of paraffin-embedded human lung carcinoma, untreated (left) or λ phosphatase-treated (right), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb.

Flow Cytometry

Flow cytometric analysis of Jurkat cells, U0126-treated (blue) or PMA-treated (green), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb.

IF-IC

Confocal immunofluorescent analysis of Drosophila egg chambers, untreated (top) or λ phosphatase-treated (bottom), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb #4370 (green) and S6 Ribosomal Protein (54D2) Mouse mAb #2317 (red). Blue pseudocolor = DRAQ5^® #4084 (fluorescent DNA dye).

IF-IC

Confocal immunofluorescent analysis of C2C12 cells, treated with U0126 #9903 (left, 10 μM for 1 h) or TPA #9905 (right, 200 nM for 15 min), using Phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) (D13.14.4E) XP™ Rabbit mAb (green). Actin filaments have been labeled with Alexa Fluor^® 555 phalloidin (red). Blue pseudocolor = DRAQ5^® #4084 (fluorescent DNA dye).

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 is 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.

Application References

Gray, M.J. et al. (2008) J Natl Cancer Inst 100, 109-20. Applications: Western Blotting
Rygiel, T.P. et al. (2008) J Cell Sci 121, 1183-92. Applications: Western Blotting
Engelman, J.A. et al. (2008) Nat Med 14, 1351-6. Applications: IHC-P (paraffin) Western Blotting
Mabilleau, G. and Sabokbar, A. (2009) PLoS ONE 4, e4173. Applications: Western Blotting
Koenigsmann, J. et al. (2009) Blood 113, 4690-701. Applications: Flow Cytometry
Schrage, Y.M. et al. (2009) Cancer Res 69, 6216-22. Applications: Western Blotting
Miyaji, M. et al. (2009) Proc Natl Acad Sci U S A 106, 14502-7. Applications: Western Blotting
Kubo, T. et al. (2009) J Exp Med 206, 1971-82. Applications: Western Blotting
Hudon, V. et al. (2010) J Med Genet 47, 182-9. Applications: IHC-P (paraffin)
Kippenberger, S. et al. (2010) Biochim Biophys Acta 1803, 940-950. Applications: IF-IC (In Cells) Western Blotting

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This product is intended for research purposes only. The product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.