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Alexa Fluor® 488 Mouse anti-MAPKAPK-2 (pT334)
Alexa Fluor® 488 Mouse anti-MAPKAPK-2 (pT334)
Analyses of MAPKAPK2 (pT334) expression by Human and Mouse Cells. Human Cells      Panel 1a: Flow Cytometric analysis of MAPKAPK2 (pT334) in 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 stained with BD Phosflow™ Alexa Fluor® 488 Mouse Anti-MAPKAPK2 (pT334) (Cat. No. 562471) 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.      Panel 1b: Western blot analysis of MAPKAPK2 (pT334) expressed by peripheral blood mononuclear cells (PBMC). Lysates from 1X10^6 untreated (C) and PMA-treated (T) PBMC were blotted using Purified Mouse Anti-MAPKAPK2 (pT334) mAb (2 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 Alexa Fluor® 488 Mouse anti-MAPKAPK2 (pT334) Ab and analyzed by flow cytometry as described above.
Analyses of MAPKAPK2 (pT334) expression by Human and Mouse Cells. Human Cells      Panel 1a: Flow Cytometric analysis of MAPKAPK2 (pT334) in 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 stained with BD Phosflow™ Alexa Fluor® 488 Mouse Anti-MAPKAPK2 (pT334) (Cat. No. 562471) 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.      Panel 1b: Western blot analysis of MAPKAPK2 (pT334) expressed by peripheral blood mononuclear cells (PBMC). Lysates from 1X10^6 untreated (C) and PMA-treated (T) PBMC were blotted using Purified Mouse Anti-MAPKAPK2 (pT334) mAb (2 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 Alexa Fluor® 488 Mouse anti-MAPKAPK2 (pT334) Ab and analyzed by flow cytometry as described above.
Product Details
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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_11154411
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 to Alexa Fluor® 488 under optimum conditions, and unreacted Alexa Fluor® 488 was removed. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.

Product Notices

  1. 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).
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. Alexa Fluor® 488 fluorochrome emission is collected at the same instrument settings as for fluorescein isothiocyanate (FITC).
  4. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  5. The Alexa Fluor®, Pacific Blue™, and Cascade Blue® dye antibody conjugates in this product are sold under license from Molecular Probes, Inc. for research use only, excluding use in combination with microarrays, or as analyte specific reagents. The Alexa Fluor® dyes (except for Alexa Fluor® 430), Pacific Blue™ dye, and Cascade Blue® dye are covered by pending and issued patents.
  6. Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
  7. 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.
  8. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  9. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
562471 Rev. 1
Antibody Details
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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.

562471 Rev. 1
Format Details
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Alexa Fluor™ 488
Alexa Fluor™ 488 Dye is part of the BD blue family of dyes. This is a small organic fluorochrome with an excitation maximum (Ex Max) at 494-nm and an emission maximum (Em Max) at 517-nm. Alexa Fluor™ 488 is designed to be excited by the Blue laser (488 nm) and detected using an optical filter centered near 520-nm (e.g., a 530/30-nm bandpass filter). Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
Alexa Fluor™ 488
Blue 488 nm
494 nm
517 nm
562471 Rev.1
Citations & References
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Development References (8)

  1. 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
  2. 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
  3. 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
  4. 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
  5. 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
  6. 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
  7. 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
  8. 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
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562471 Rev. 1

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For Research Use Only. Not for use in diagnostic or therapeutic procedures.