Alexa Fluor® 488 Mouse anti-MEK1 (pS298)

BD Phosflow™ Alexa Fluor® 488 Mouse anti-MEK1 (pS298)

Name: Alexa Fluor® 488 Mouse anti-MEK1 (pS298)
Brand: BD Phosflow™
SKU: 560192

Clone J114-64

(RUO)

Analysis of MEK1 (pS298) in human epithelioid carcinoma. Serum-starved HeLa S3 cells (ATCC CCL 2.2) were either stimulated (shaded histogram) with 50 nM Calyculin A (Calbiochem, Cat. No. 208851) at 37˚C for 30 minutes or unstimulated (open histogram). The cells were fixed (BD Cytofix™ buffer, Cat. No. 554655) for 10 minutes at 37°C, then permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for at least 30 minutes, and then stained with Alexa Fluor® 488 Mouse anti-MEK1 (pS298, Cat. No. 560192). Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.

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Product Details

Brand: BD Phosflow™

Alternative Name: MAP2K1, MAPKK1, MKK1, MP2K1, PPKMK1

Reactivity: Human (QC Testing), Mouse (Reactivity Confirmed in Development), Rat (Predicted)

Isotype: Mouse BALB/c IgG1, κ

Immunogen: Phosphorylated Human MEK1 Peptide

Application: Intracellular staining (flow cytometry) (Routinely Tested)

Vol. Per Test: 20 µl

RRID: AB_1645360

Storage Buffer: Aqueous buffered solution containing BSA and ≤0.09% sodium azide.

Regulatory Status: RUO

Preparation And Storage

Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to Alexa Fluor® 488 under optimum conditions, and unreacted Alexa Fluor® 488 was removed.

Recommended Assay Procedures

Either BD Cytofix™ fixation buffer or BD Phosflow™ Fix Buffer I may be used for cell fixation.

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).
Source of all serum proteins is from USDA inspected abattoirs located in the United States.
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.
Alexa Fluor® 488 fluorochrome emission is collected at the same instrument settings as for fluorescein isothiocyanate (FITC).
For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
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.
Alexa Fluor® is a registered trademark of Molecular Probes, Inc., Eugene, OR.
Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.

Companion Products

Stain Buffer (FBS) RUO

Size 500 mL Cat No. 554656

Stain Buffer (BSA) RUO

Size 500 mL Cat No. 554657

Fixation Buffer RUO

Size 100 mL Cat No. 554655

Fix Buffer I RUO

Size 250 mL Cat No. 557870

Perm Buffer III RUO

Size 125 mL Cat No. 558050

560192 Rev. 2

Antibody Details

J114-64

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 by activated MAP kinase kinase kinases (MAP3Ks), such as the Raf isoforms. Hormones, growth and differentiating factors, or tumor promoters induce Raf activation via activation of Ras proteins. Alternatively, cellular adhesion can lead to phosphorylation of MEK1 at serine 298 (S298), mediated by p21-activated kinase (PAK). The S298-phosphorylated MEK1 has an enhanced capacity to interact with Raf, resulting in MEK1 activation.

The J114-64 monoclonal antibody recognizes the phosphorylated S298 of MEK1.

560192 Rev. 2

Format Details

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.

Format: Alexa Fluor™ 488

Excitation Source: Blue 488 nm

Excitation Max: 494 nm

Emission Max: 517 nm

560192 Rev.2

Citations & References

Development References (3)

Eblen ST, Slack JK, Weber MJ, Catling AD. Rac-PAK signaling stimulates extracellular signal-regulated kinase (ERK) activation by regulating formation of MEK1-ERK complexes. Mol Cell Biol. 2002; 22(17):6023-6033. (Biology).
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
Slack-Davis JK, Eblen ST, Zecevic M, et al. PAK1 phosphorylation of MEK1 regulates fibronectin-stimulated MAPK activation. J Cell Biol. 2003; 162(2):281-291. (Biology).

560192 Rev. 2

Please refer to Support Documents for Quality Certificates

Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described

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.