Analyses of MAPKAPK2 (pT334) expression by Human and Mouse Cells. Human Cells Panel 1a: Flow Cytometric analysis of MAPKAPK2 (pT334) expressed by peripheral blood lymphocytes (PBL). Whole blood cells were not stimulated (dashed line histogram) or stimulated (solid line histogram) with 400 nM Phorbol 12-Myristate 13-Acetate (PMA; Sigma, Cat. No. P8139; 15 min, 37°C). Cells were fixed in 1X BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049; 10 min, 37˚C) and permeabilized in BD Phosflow™ Perm Buffer III (Cat. No. 558050) on ice (30 min). Cells were then stained with BD Phosflow™ PE Mouse Anti-MAPKAPK2 (pT334) (Cat. No. 562470) antibody. Fluorescence histograms showing MAPKAPK2 (pT334) expression were generated for gated events with the light scatter characteristics of intact lymphocytes using a BD FACSCanto™ II Flow Cytometer System. Panel 1b: Western blot analysis of MAPKAPK2 (pT334) expressed by human peripheral blood mononuclear cells (PBMC). Lysates from 1X10^6 untreated (C) and PMA-treated (T) PBMC were blotted using Purified Mouse Anti-MAPKAPK2 (pT334) antibody (2.0 µg/ml, Cat. No. 562469), HRP Goat Anti-Mouse Ig (Cat. No. 554002) and a chemiluminescent detection system. MAPKAPK2 (pT334) were identified as ~49 kDa bands by Western blotting. Mouse Cells Panel 2: Splenocytes were not stimulated (dashed line histogram) or stimulated (solid line histogram) with PMA (50 nM, 15 min, 37°C). Cells were fixed, permeabilized, stained with PE Mouse anti-MAPKAPK2 (pT334) antibody and analyzed by flow cytometry as above.
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Analyses of MAPKAPK2 (pT334) expression by Human and Mouse Cells. Human Cells Panel 1a: Flow Cytometric analysis of MAPKAPK2 (pT334) expressed by peripheral blood lymphocytes (PBL). Whole blood cells were not stimulated (dashed line histogram) or stimulated (solid line histogram) with 400 nM Phorbol 12-Myristate 13-Acetate (PMA; Sigma, Cat. No. P8139; 15 min, 37°C). Cells were fixed in 1X BD Phosflow™ Lyse/Fix Buffer (Cat. No. 558049; 10 min, 37˚C) and permeabilized in BD Phosflow™ Perm Buffer III (Cat. No. 558050) on ice (30 min). Cells were then stained with BD Phosflow™ PE Mouse Anti-MAPKAPK2 (pT334) (Cat. No. 562470) antibody. Fluorescence histograms showing MAPKAPK2 (pT334) expression were generated for gated events with the light scatter characteristics of intact lymphocytes using a BD FACSCanto™ II Flow Cytometer System. Panel 1b: Western blot analysis of MAPKAPK2 (pT334) expressed by human peripheral blood mononuclear cells (PBMC). Lysates from 1X10^6 untreated (C) and PMA-treated (T) PBMC were blotted using Purified Mouse Anti-MAPKAPK2 (pT334) antibody (2.0 µg/ml, Cat. No. 562469), HRP Goat Anti-Mouse Ig (Cat. No. 554002) and a chemiluminescent detection system. MAPKAPK2 (pT334) were identified as ~49 kDa bands by Western blotting. Mouse Cells Panel 2: Splenocytes were not stimulated (dashed line histogram) or stimulated (solid line histogram) with PMA (50 nM, 15 min, 37°C). Cells were fixed, permeabilized, stained with PE Mouse anti-MAPKAPK2 (pT334) antibody and analyzed by flow cytometry as above.
Product Details
BD Phosflow™
MAP kinase-activated protein kinase 2, MAPKAP kinase 2; MK2; MAPK2
Human (QC Testing), Mouse (Tested in Development)
Mouse IgG1, κ
Phosphorylated Human MAPKAPK-2 Peptide
Intracellular staining (flow cytometry) (Routinely Tested)
5 µl
AB_11154235
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO
Preparation And Storage
Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The antibody was conjugated with R-PE under optimum conditions, and unconjugated antibody and free PE were removed. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.
Product Notices
- This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
- An isotype control should be used at the same concentration as the antibody of interest.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- 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.
- Source of all serum proteins is from USDA inspected abattoirs located in the United States.
Companion Products
Stain Buffer (FBS) RUO
Size
500 mL
Cat No.
554656
Lyse/Fix Buffer 5X RUO
Size
250 mL
Cat No.
558049
Perm/Wash Buffer I RUO
Size
125 mL
Cat No.
557885
Perm Buffer II RUO
Size
125 mL
Cat No.
558052
Perm Buffer III RUO
Size
125 mL
Cat No.
558050
Perm Buffer IV 10× RUO
Size
50 mL
Cat No.
560746
562470 Rev. 1
Antibody Details
P24-694
The P24-694 monoclonal antibody specifically binds to the phosphorylated T334 site (pT334) of MAPKAPK-2. MAPKAPK-2 is a serine/threonine protein kinase. This ~49 kDa member of the MAPKAPK family of protein kinases is also known as mitogen-activated protein kinase-activated protein kinase 2. MAPKAPK-2 is phosphorylated and activated by p38 MAP kinase in response to stress, cytokines and chemokines. MAPKAPK-2 is phosphorylated on multiple sites including Thr222, Ser272 and Thr334. Phosphorylation of any two of these three amino acid residues seems to be required for the activation of this kinase that serves multiple cellular functions. Phosphorylation of Thr334 was reported to be essential for nuclear export of the heterodimer formed between p38 MAPK and MAPKAPK-2. Mice deficient in MAPKAPK-2 have been shown to be protected from ischemic injury. MAPKAPK-2 is also reported to serve as a cell cycle checkpoint kinase in response to UV irradiation. The heat shock protein, HSP27 was shown to be one of the major substrates of MAPK and MAPKAPK-2.
562470 Rev. 1
Format Details
PE
R-Phycoerythrin (PE), is part of the BD family of Phycobiliprotein dyes. This fluorochrome is a multimeric fluorescent phycobiliprotein with excitation maximum (Ex Max) of 496 nm and 566 nm and an emission maximum (Em Max) at 576 nm. PE is designed to be excited by the Blue (488 nm), Green (532 nm) and Yellow-Green (561 nm) lasers and detected using an optical filter centered near 575 nm (e.g., a 575/26-nm bandpass filter). As PE is excited by multiple lasers, this can result in cross-laser excitation and fluorescence spillover on instruments with various combinations of Blue, Green, and Yellow-Green lasers. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
PE
Yellow-Green 488 nm, 532 nm, 561 nm
496 nm, 566 nm
576 nm
562470 Rev.1
Citations & References
View product citations for antibody "562470" on CiteAb
Development References (9)
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Ben-Levy R, Leighton IA, Doza YN, et al. Identification of novel phosphorylation sites required for activation of MAPKAP kinase-2. EMBO J. 14(23):5920-5930. (Biology). View Reference
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Clifton AD, Young PR, Cohen P. A comparison of the substrate specificity of MAPKAP kinase-2 and MAPKAP kinase-3 and their activation by cytokines and cellular stress. FEBS Lett. 1996; 392(3):209-214. (Biology). View Reference
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Engel K, Kotlyarov A, Gaestel M. Leptomycin B-sensitive nuclear export ofMAPKAP kinase 2 is regulated by phosphorylation. EMBO J. 1998; 17(12):3363. (Biology). View Reference
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Heidenreich O, Neininger A, Schratt G, et al. MAPKAP kinase 2 phosphorylates serum response factor in vitro and in vivo. J Biol Chem. 1999; 274(20):14434-14443. (Biology). View Reference
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Krump E, Sanghera JS, Pelech SL, Furuya W, Grinstein S. Chemotactic peptideN-formyl-met-leu-phe activation of p38 mitogen-activated protein kinase (MAPK)and MAPK-activated protein kinase-2 in human neutrophils. J Biol Chem. 1997; 272(2):937. (Biology). View Reference
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Manke IA, Nguyen A, Lim D, Stewart MQ, Elia AE, Yaffe MB. MAPKAP kinase-2 is acell cycle checkpoint kinase that regulates the G2/M transition and S phaseprogression in response to UV irradiation. Mol Cell. 2005; 17(1):37. (Biology). View Reference
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Meng W, Swenson LL, Fitzgibbon MJ, et al. Structure of mitogen-activated protein kinase-activated protein (MAPKAP) kinase 2 suggests a bifunctional switch that couples kinase activation with nuclear export. J Biol Chem. 2002; 277(40):37401-37405. (Biology). View Reference
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Rouse J, Cohen P, Trigon S, et al. A novel kinase cascade triggered by stress and heat shock that stimulates MAPKAP kinase-2 and phosphorylation of the small heat shock proteins. Cell. 1994; 78(6):1027-1037. (Biology). View Reference
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Wang X, Xu L, Wang H, Young PR, Gaestel M, Feuerstein GZ.. Mitogen-activatedprotein kinase-activated protein (MAPKAP) kinase 2 deficiency protects brain fromischemic injury in mice. J Biol Chem. 2002; 277(46):43968. (Biology). View Reference
562470 Rev. 1
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For Research Use Only. Not for use in diagnostic or therapeutic procedures.