Invitrogen

Phospho-ERK1/ERK2 (Thr185, Tyr187) Polyclonal Antibody

Advanced Verification

This Antibody was verified by Cell Treatment to ensure that the antibody binds to the antigen stated. View Details

18 Published Figures

45 References

View all (49) ERK1/2 antibodies

Datasheet

Datasheet

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Phospho-ERK1/ERK2 (Thr185, Tyr187) Antibody — Figure 4 Haem-dependent dimerization of PGRMC1 is necessary for tumour proliferation mediated by EGFR signalling. ( a ) FLAG-PGRMC1 wild-type (wt) and Y113F mutant proteins (a.a.44-195), in either apo- or haem-bound form, were incubated with purified EGFR and co-immunoprecipitated with anti-FLAG antibody-conjugated beads. Input and bound proteins were detected by Western blotting. ( b ) In vitro binding assay was performed as in ( a ) using haem-bound FLAG-PGRMC1 wt (a.a.44-195) and purified EGFR with or without treatment of RuCl 3 and CORM3. ( c ) FLAG-PGRMC1 wt or Y113F (full length) was over-expressed in HCT116 cells and immunoprecipitated with anti-FLAG antibody-conjugated beads. Co-immunoprecipitated proteins (FLAG-PGRMC1, endogenous PGRMC1 and EGFR) were detected with Western blotting by using anti-PGRMC1 or anti-EGFR antibody. ( d ) HCT116 cells were treated with or without 250 mumol l -1 of succinylacetone (SA) for 48 h. The intracellular haem was extracted and quantified by reverse-phase HPLC. The data represent mean+-s.d. of four separate experiments. ** P <0.01 using unpaired Student and apos;s t -test. ( e ) Co-immunoprecipitation assay was performed as in ( c ) with or without SA treatment in HCT116 cells. ( f ) HCT116 cells expressing control shRNA or those knocking down PGRMC1 (PGRMC1-KD) were treated with EGF or left untreated, and components of the EGFR signaling pathway were detected by Western blotting. ( g , h ) HCT116 cells were treated with or without EGF, S

Phospho-ERK1/ERK2 (Thr185, Tyr187) Antibody in Immunohistochemistry (Paraffin) (IHC (P)) — Immunohistochemistry analysis of ERK1/2 (pTpY185/187) showing staining in the cytoplasm and nucleus of paraffin-embedded human breast carcinoma tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a ERK1/2 (pTpY185/187) polyclonal antibody (Product # 44-680G) diluted in 3% BSA-PBS at a dilution of 1:50 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Phospho-ERK1/ERK2 (Thr185, Tyr187) Antibody in Western Blot (WB) — Western blot analysis was performed on whole cell extracts (30 µg lysate) of MDA-MB-231 with treatment of EGF(100 ng/mL for 15 mins) (Lane 1), MDA-MB-231 (Lane 2), U-87 MG (Lane 3),, SH-SY5Y (Lane 4) and HeLa (Lane 5). The blots were probed with Anti-Phospho-p44 MAPK + p42 MAPK pThr185 + pTyr187 Rabbit polyclonal Antibody (Product# 44680G, 1 in 1000 dilution) and detected by chemiluminescence Goat anti-Rabbit IgG (H+L) Secondary Antibody, HRP conjugate (Product # G21234, 1:5000 dilution). Bands of 42 kDa and 44 kDa corresponding to Phospho-p44 MAPK + p42 MAPK pThr185 + pTyr187 was observed across cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 12 % Bis-Tris gel (Product # NP0341BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Phospho-ERK1/ERK2 (Thr185, Tyr187) Antibody in Cell Treatment — Altered expression of proteins upon cell treatment demonstrates antibody specificity. Western blot of Phospho-ERK1/ERK2 (Thr185, Tyr187) using Anti - ERK1/2 [pTpY185/187]Rabbit polyclonal Antibody (Product # 44-680G), shows increase in expression of Phospho-ERK1/ERK2 (Thr185, Tyr187upon treatment with EGF in MDA-MB-231 cells. {TM}

Product Details

Tested Applications

Dilution

Immunohistochemistry (Paraffin) (IHC (P))

1:10-1:100

Western Blot (WB)

1:1000

Published Applications

Western Blot (WB)

See 29 publications below

Miscellaneous PubMed (MISC)

See 7 publications below

Immunofluorescence (IF)

See 2 publications below

Immunocytochemistry (ICC)

See 3 publications below

Immunohistochemistry (Paraffin) (IHC (P))

See 2 publications below

Immunohistochemistry (IHC)

See 1 publication below

Immunohistochemistry - Free Floating (IHC (Free))

See 1 publication below

Product Specifications

Species Reactivity

C. elegans, Chicken, Fruit fly, Human, Mouse, Rat, Xenopus, Bovine

Published species

C. elegans, Human, Mouse, Rat

Host / Isotype

Rabbit / IgG

Class

Polyclonal

Type

Antibody

Immunogen

The antiserum was produced against a chemically synthesized phosphopeptide derived from the region of human ERK1 and 2 that contains threonine 202/185 and tyrosine 204/187. This region is conserved among many species including rat, mouse, cow, frog, snail, nematode, and fruit fly.

Conjugate

Unconjugated

Form

Liquid

Purification

Antigen affinity chromatography

Storage buffer

Dulbecco's PBS, pH 7.3, with 50% glycerol, 1mg/mL BSA

Contains

0.05% sodium azide

Storage conditions

-20°C

RRID

AB_2533719

Product Specific Information

Purified from rabbit serum by sequential epitope-specific chromatography, this product contains enough material for 10 mini-blots. The antibody has been negatively preadsorbed using a non-phosphopeptide corresponding to the sites of phosphorylation to remove antibody that is reactive with non-phosphorylated ERK1 and 2. The final product is generated by affinity chromatography using an ERK1 and 2-derived peptide that is phosphorylated at threonine 202/185 and tyrosine 204/187, respectively, within the activation loop.

This antibody has been used in western blotting and previous lots have been used in immunostaining. Other applications may work but have not been tested. The positive control used in western analysis were PC12 cells +/- Sorbitol; NIH3T3 +/- PDGF; and HEK293 +/- UV.

Target Information

ERK (extracellular signal regulated kinase), also known as MAPK (mitogen-activated protein kinase) has two closely related isoforms designated ERK1 and ERK2 (44 kDa and 42 kDa, respectively). These kinases belong to a family of serine/threonine kinases that are activated upon treatment of cells with a large variety of stimuli including mitogens, hormones, growth factors, cytokines, virus infection, ligands for heterotrimeric guanine nucleotide-binding protein (G protein)-coupled receptors, transforming agents, and bioactive peptides. ERK1 and ERK2 are widely expressed and are involved in the regulation of meiosis, mitosis, and postmitotic functions in differentiated cells. Cell stimulation induces the activation of a signaling cascade, the downstream effects of which have been linked to the regulation of cell growth and differentiation as well as the cytoskeleton. ERK1 and ERK2 are phosphorylated within the activation loop on both a threonine and a tyrosine residue (within a Thr-Glu-Tyr motif) by MEKs (MAPK/ERK kinases), thereby greatly elevating the activity of ERK1/2. When growth factors bind to their receptor tyrosine kinase, Ras interacts with Raf, the serine/threonine protein kinase, and activates it, as well. Once activated, Raf phosphorylates serine residue in 2 additional kinases, MEK1/2, which in turn phosphorylates tyrosine/threonine in ERK1/2. Upon activation, the ERKs either phosphorylate a number of cytoplasmic targets or migrate to the nucleus, where they phosphorylate and activate a number of transcription factors such as c-Fos and Elk-1.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

Bioinformatics

Protein Aliases: ERK-1; ERK-2; ERK1; ERK2; ERT1; ERT2; Extracellular signal-regulated kinase 1; Extracellular signal-regulated kinase 2; extracellular signal-related kinase 1; extracellular-signal-regulated kinase 2; Insulin-stimulated MAP2 kinase; M phase MAP kinase; MAP kinase; MAP kinase 1; MAP kinase 2; MAP kinase 3; MAP kinase isoform p42; MAP kinase isoform p44; MAPK 1; MAPK 2; MAPK 3; MAPK1; MAPK3; MBP kinase; Microtubule-associated protein 2 kinase; mitogen activated protein kinase 1; mitogen-activated 3; Mitogen-activated protein kinase 1; mitogen-activated protein kinase 1a; Mitogen-activated protein kinase 2; Mitogen-activated protein kinase 3; MNK1; Myelin basic protein kinase; Myelin xP42 protein kinase; p42-MAPK; p44 MAP kinase; p44-ERK1; p44-MAPK; pp42/MAP kinase; PRKM1; PRKM2; PRKM3; protein tyrosine kinase ERK2

Gene Aliases: 9030612K14Rik; AA407128; AU018647; BOS_16809; BOS_22892; C78273; ERK; ERK-1; ERK-2; ERK1; ERK2; ERT1; ERT2; Esrk1; HS44KDAP; HUMKER1A; Mapk; MAPK1; mapk1-a; mapk1-b; mapk1a; MAPK2; MAPK3; MNK1; mpk1; Mtap2k; p38; p40; p41; p41mapk; p42-MAPK; P42MAPK; p44; p44-ERK1; p44-MAPK; P44ERK1; P44MAPK; PRKM1; PRKM2; PRKM3; XELAEV_18010534mg; xp42

UniProt ID: (Bovine) P46196, (Human) P28482, (Human) P27361, (Mouse) P63085, (Rat) P63086, (Rat) P21708, (Mouse) Q63844, (Xenopus) P26696

Entrez Gene ID: (Bovine) 327672, (Chicken) 373953, (Human) 5594, (Bovine) 531391, (Human) 5595, (Mouse) 26413, (Rat) 116590, (Rat) 50689, (Mouse) 26417, (Xenopus) 398985