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Purified Rabbit Anti-MEK1/2 (pS222)
Purified Rabbit Anti-MEK1/2 (pS222)
Analysis of MEK1/2 (pS222) in activated human lymphocytes.  Peripheral blood mononuclear cells were either stimulated with PMA and Ionomycin at 37˚C for 10 minutes or unstimulated.  The cells were fixed with BD Cytofix™ buffer (Cat. No. 554655), permeabilized in BD Phosflow Perm Buffer III (Cat. No. 558050), and stained with Purified Rabbit anti-MEK1/2 (pS222) followed by PE F(ab')2 Donkey anti-Rabbit IgG (Multiple Adsorption, Cat. No. 558416).  Flow cytometry was performed on a BD FACSCalibur flow cytometry system; the lymphocytes were selected according to their scatter profile.  The open and gray-filled histograms display MEK1/2 (pS222) expression on activated and unactivated lymphocytes, respectively, while the black-filled histogram shows activated lymphocytes stained with the PE second-step antibody alone.
Purified Rabbit Anti-MEK1/2 (pS222)

MEK1/2 (pS222) staining on tonsil.  Fresh human tonsil was incubated in 5 mM Pervanadate solution for 2 hours (top row) or untreated (bottom row), then fixed in formalin and processed.  Following antigen retrieval with BD Retrievagen A buffer (Cat. no. 550524), the sections were either left untreated (left column) or treated with a phosphatase to eliminate all phosphorylation (right column).  The tissue sections were stained with affinity purified Rabbit anti-MEK1/2 (pS222) with Hematoxylin counterstaining.  Original magnification: 40X.

Analysis of MEK1/2 (pS222) in activated human lymphocytes.  Peripheral blood mononuclear cells were either stimulated with PMA and Ionomycin at 37˚C for 10 minutes or unstimulated.  The cells were fixed with BD Cytofix™ buffer (Cat. No. 554655), permeabilized in BD Phosflow Perm Buffer III (Cat. No. 558050), and stained with Purified Rabbit anti-MEK1/2 (pS222) followed by PE F(ab')2 Donkey anti-Rabbit IgG (Multiple Adsorption, Cat. No. 558416).  Flow cytometry was performed on a BD FACSCalibur flow cytometry system; the lymphocytes were selected according to their scatter profile.  The open and gray-filled histograms display MEK1/2 (pS222) expression on activated and unactivated lymphocytes, respectively, while the black-filled histogram shows activated lymphocytes stained with the PE second-step antibody alone.

MEK1/2 (pS222) staining on tonsil.  Fresh human tonsil was incubated in 5 mM Pervanadate solution for 2 hours (top row) or untreated (bottom row), then fixed in formalin and processed.  Following antigen retrieval with BD Retrievagen A buffer (Cat. no. 550524), the sections were either left untreated (left column) or treated with a phosphatase to eliminate all phosphorylation (right column).  The tissue sections were stained with affinity purified Rabbit anti-MEK1/2 (pS222) with Hematoxylin counterstaining.  Original magnification: 40X.

Product Details
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BD Phosflow™
Human (QC Testing), Mouse, Rat (Reported)
Rabbit IgG
Phosphorylated Human MEK1 and MEK2 peptide
Intracellular staining (flow cytometry) (Routinely Tested), Immunohistochemistry-formalin (antigen retrieval required), Western blot (Tested During Development)
20 µl
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO


Preparation And Storage

The polyclonal antibody was purified from antiserum by negative adsorption and affinity chromatography. Store undiluted at 4°C.

Recommended Assay Procedures

This purified polyclonal antibody is suitable for intracellular staining of human whole blood (using BD™ Phosflow Lyse/Fix Buffer) and peripheral blood mononuclear cells (using BD Cytofix™ Fixation Buffer).  

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. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  3. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  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.
558281 Rev. 3
Antibody Details
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poly/MEK1/2/44452

MEK (Map/Erk Kinase) 1 and 2 are serine/threonine kinases, also known as MAP kinase kinases (MAP2K1 and 2, MAPKK1 and 2, or MKK1 and 2).  They activate the MAP (Mitogen-Activated Protein) kinases, also known as ERKs (Extracellular signal Regulated Kinases), which are critical kinases in multiple signal transduction pathways that regulate cell growth and differentiation.  Activation of MEK 1 and 2 is dependent upon phosphorylation of serines 218 and/or 222 (S222) by MAP kinase kinase kinases (MAP3Ks), such as the Raf isoforms.

The affinity purified rabbit anti-MEK1/2 (pS222) polyclonal antibody recognizes human MEK1 and 2 phosphorylated at S222 in flow cytometric analysis of human blood cells.  The same polyclonal antibody preparation is able to identify human, mouse, and rat MEK1/2 (pS222) in western blot analysis.  The sequence is conserved in several other species, including chimpanzee, chicken, and frog.

558281 Rev. 3
Format Details
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Purified
Tissue culture supernatant is purified by either protein A/G or affinity purification methods. Both methods yield antibody in solution that is free of most other soluble proteins, lipids, etc. This format provides pure antibody that is suitable for a number of downstream applications including: secondary labeling for flow cytometry or microscopy, ELISA, Western blot, etc.
Purified
558281 Rev.3
Citations & References
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Development References (3)

  1. Crews CM, Alessandrini A, Erikson RL. The primary structure of MEK, a protein kinase that phosphorylates the ERK gene product. Science. 1992; 258(5081):478-480. (Biology). View Reference
  2. Kolch W. Meaningful relationships: the regulation of the Ras/Raf/MEK/ERK pathway by protein interactions. Biochem J. 2000; 351:289-305. (Biology). View Reference
  3. Papin C, Eychene A, Brunet A, et al. B-Raf protein isoforms interact with and phosphorylate Mek-1 at serine residues 218 and 222. Oncogene. 1995; 10(8):1647-1651. (Biology). View Reference
558281 Rev. 3

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Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

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