Mitogen-Activated Protein Kinase 14 (MAPK14) (pThr180,pTyr182) antibody

Antigen: Mitogen-Activated Protein Kinase 14 (MAPK14)

Synonyms: RK, p38, EXIP, Mxi2, CSBP1, CSBP2, CSPB1, PRKM14, PRKM15, SAPK2A, p38ALPHA, Hog, CRK1, CSBP, Exip, Csbp1, Csbp2, Prkm14, Prkm15, Sapk2A, p38Hog, p38alpha, MGC105413, p38b, zp38b, MAPK14, MGC142910, 186F5S, BG:DS00797.3, D-p38, D-p38b, Dm p38b, Dp38, ESTS:186F5S, Mpk34C, anon-sts23, p38 MAPK, p38 beta, p38B, p38beta, DmelCG7393, CG7393

Binding Site: pThr180,pTyr182

Clonality: Monoclonal (36)

Host: Mouse

Reactivity: Human

Conjugate: Un-conjugated

Application: Western Blotting (WB), BioImaging (BI), Intracellular Flow Cytometry (ICFC)

Catalog no.: ABIN968803

Additional Information

Alternative name: p38 MAPK

Immunogen: Phosphorylated Human p38 MAPK (pT180/pY182) Peptide

Cross-Reactivity: Mouse (Murine), Rat (Rattus)

Format: Liquid

Isotype: IgG1

Clone: 36

Description: Activation of the immune and inflammatory responses often involves the recognition of bacterial endotoxin (lipopolysaccharide or LPS). Binding of LPS by monocytes results in the production and release of proinflammatory cytokines, such as IL-1 and TNF. LPS-induced signaling cascades involve members of the Ser/Thr protein kinase family known as the Mitogen Activated Protein Kinases (MAPKs). MAPK signal transduction pathways mediate the effects of various extracellular stimuli on biological processes such as proliferation, differentiation, and death. The p38 MAPKs include p38alpha (MAPK14), beta (MAPK11), gamma (MAPK12), and delta (MAPK13). These Ser/Thr kinases are activated by dual phosphorylation on threonine (T) and tyrosine (Y) within the motif Thr-Gly-Tyr located in kinase subdomain VIII. Activation of p38 MAPK is mediated specifically by the MAP Kinase Kinases, MKK3, MKK4, and MKK6. This leads to the activation of multiple transcription factors (NF-kappaB, ATF-2, Elk-1, and CHOP) that induce expression of many different genes, including proinflammatory cytokine genes. Thus, p38 MAPKs are central kinases in multiple signal transduction pathways.
The 36/p38 (pT180/pY182) monoclonal antibody recognizes the conserved dual phosphorylated site pT180/pY182 of p38alpha, beta, gamma, and delta.
Synonyms: MK14, 11, 12, 13, CSBP1, SAPK2, 2A, 3, 4, MX12, ERK-6, ERK5

Characteristics: 1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
2. Please refer to us for technical protocols.
3. This antibody has been developed and certified for the bioimaging application. However, a routine bioimaging test is not performed on every lot. Researchers are encouraged to titrate the reagent for optimal performance.
4. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
5. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
6. Triton is a trademark of the Dow Chemical Company.

Molecular Weight: 38-42 kDa

Comments:

Related Products: ABIN968535, ABIN968632, ABIN968769

Synonyms: RK, p38, EXIP, Mxi2, CSBP1, CSBP2, CSPB1, PRKM14, PRKM15, SAPK2A, p38ALPHA, Hog, CRK1, CSBP, Exip, Csbp1, Csbp2, Prkm14, Prkm15, Sapk2A, p38Hog, p38alpha, MGC105413, p38b, zp38b, MAPK14, MGC142910, 186F5S, BG:DS00797.3, D-p38, D-p38b, Dm p38b, Dp38, ESTS:186F5S, Mpk34C, anon-sts23, p38 MAPK, p38 beta, p38B, p38beta, DmelCG7393, CG7393

Application Details

Application Notes: Bioimaging
1. Seed the cells in appropriate culture medium at ~10,000 cells per well in an 96-well Imaging Plate and culture overnight.
2. Remove the culture medium from the wells, and fix the cells by adding 100 myl of Fixation Buffer to each well. Incubate for 10 minutes at room temperature (RT).
3. Remove the fixative from the wells, and permeabilize the cells using either 90% methanol, or Triton™ X-100: a. Add 100 myl of -20°C 90% methanol to each well and incubate for 5 minutes at RT. OR b. Add 100 myl of 0.1% Triton™ X-100 to each well and incubate for 5 minutes at RT.
4. Remove the permeabilization buffer, and wash the wells twice with 100 myl of 1× PBS.
5. Remove the PBS, and block the cells by adding 100 myl of to each well. Incubate for 30 min. at RT.
6. Remove the blocking buffer and add 50 myl of the optimally titrated primary antibody (diluted in Stain Buffer) to each well, and incubate 1 hr.
7. Remove the primary antibody, and wash the wells three times with 100 myl of 1× PBS.
8. Remove the PBS, and add the second step reagent at its optimally titrated concentration in 50 myl to each well, and incubate in the dark for 1 hr.
9. Remove the second step reagent, and wash the wells three times with 100 myl of 1× PBS.
10. Remove the PBS, and counter-stain the nuclei by adding 200 myl per well of 2 myg/ml Hoechst 33342 in 1× PBS to each well at least 15 minutes before imaging.
11. View and analyze the cells on an appropriate imaging instrument.

Concentration: 250 µg/ml

Purity: Purified

Purification: Purified from tissue culture supernatant or ascites by affinity chromatography.

Buffer: Aqueous buffered solution containing BSA, glycerol.

Preservative: 0.09% Sodium azide.

Storage: Store undiluted at -20° C.

Research Area: Kinases/Phosphatases

Restrictions: For Research Use only

Images

Western blot analysis for p38 MAPK (pT180/pY182). HeLa cells (Human cervical epitheloid carcinoma, ATCC CCL-2) were either left untreated (lane 1) or treated (lane 2) with 25 µg/ml anisomycin, an antibiotic and protein synthesis inhibitor known to activate signal transduction pathways, for 15 minutes at 37°C. The top panel was probed with a mouse anti-p38alpha antibody and the bottom was probed with the mouse anti-p38 MAPK (pT180/pY182) antibody at a 1:2500 dilution. The target band in each panel is observable at 38-42 kD.

Immunofluorescent staining of HeLa cells. HeLa cells (Human cervical epitheloid carcinoma, ATCC CCL-2) were seeded in a 96-well imaging plate at ~ 10,000 cells per well. After overnight incubation, cells were either left untreated (left image) or exposed to anisomycin for 30 minutes (right image). After treatment cells were stained using the alcohol perm protocol and the mouse anti-p38 MAPK (pT180/pY182) antibody. The second step reagent was Alexa-Fluor® 488 goat anti-mouse IgG (Invitrogen). Phospho p38 staining is pseudo-colored green, nuclei were stained with Hoechst 33342 and are pseudo-colored blue. Note the lack of staining for phospho-p38 in the untreated cells (left) and the nuclear/cytoplasmic staining in the stimulated cells (right). The images were captured using a 20x objective. This antibody also stained U-2 OS (ATCC HTB-96) cells and can be used with either the Triton™ X-100 or alcohol perm protocols.

Cells exposed to anisomycin for 30 minutes

Publications

Product: Han, Lee, Bibbs et al.: "A MAP kinase targeted by endotoxin and hyperosmolarity in mammalian cells." in: Science (New York, N.Y.), Vol. 265, Issue 5173, pp. 808-11, 1994 (PubMed).

Brunet, Pouysségur: "Identification of MAP kinase domains by redirecting stress signals into growth factor responses." in: Science (New York, N.Y.), Vol. 272, Issue 5268, pp. 1652-5, 1996 (PubMed).

Winston, Chan, Johnson et al.: "Activation of p38mapk, MKK3, and MKK4 by TNF-alpha in mouse bone marrow-derived macrophages." in: Journal of immunology (Baltimore, Md. : 1950), Vol. 159, Issue 9, pp. 4491-7, 1997 (PubMed).