PE Mouse Anti-Stat5 (pY694)

BD Phosflow™ PE Mouse Anti-Stat5 (pY694)

Name: PE Mouse Anti-Stat5 (pY694)
Brand: BD Phosflow™
SKU: 562077

Clone 47/Stat5(pY694)

(RUO)

Analysis of Stat5 (pY694) in human lysed whole blood. Whole blood was either left untreated (unshaded) or treated with 100 ng/mL recombinant human IL-2 (Cat. No. 554603) for 15 minutes at 37°C (shaded). The samples were lysed and fixed with 1X BD Phosflow™ Lyse/Fix buffer (Cat. No. 558049) for 10 minutes at 37°C, permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for 30 minutes and were then stained with PE mouse anti-Stat5 (pY694). For data analysis, lymphocytes were selected by scatter profile. Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.

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Product Details

Brand: BD Phosflow™

Alternative Name: Signal transducer and activator of transcription 5; MGF; MPF

Reactivity: Human (QC Testing), Mouse,Rat,Sheep (Predicted)

Isotype: Mouse IgG1, κ

Immunogen: Phosphorylated Human Phosphorylated Stat5 Peptide

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

Vol. Per Test: 5 µl

RRID: AB_10894188

Storage Buffer: Aqueous buffered solution containing BSA, protein stabilizer, 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 with R-PE under optimum conditions, and unconjugated antibody and free PE were removed.

Recommended Assay Procedures

This antibody conjugate 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 or BD Phosflow™ Fix Buffer I).

This mAb was characterized by flow cytometry (Flow) and western blot analysis (WB) using these model systems:

Method Species Cells Treatment Fixation Perm buffer Result

Flow Human PBMC IL-2 Fixation Buffer III Positive Staining

Flow Human PBMC IL-2 Fixation Buffer I or II Unsatisfactory

Flow Human Whole Blood IL-2 Lyse/Fix III Positive Staining

Flow Human Whole Blood IL-2 Lyse/Fix I or II Unsatisfactory

Flow Human TF-1 cells GM-CSF Fixation Buffer III Positive Staining

Flow Human TF-1 cells GM-CSF Fixation Buffer I or II Unsatisfactory

WB Human A431 Cell Lysate EGF Not Applicable Not Applicable 92 kDa

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.
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.

Companion Products

Lyse/Fix Buffer 5X RUO

Size 250 mL Cat No. 558049

Perm Buffer III RUO

Size 125 mL Cat No. 558050

Fix Buffer I RUO

Size 250 mL Cat No. 557870

Fixation Buffer RUO

Size 100 mL Cat No. 554655

Recombinant Human IL-2 RUO

Size 10 µg Cat No. 554603

Purified Mouse Anti-Human Stat5 (pY694) RUO

Size 50 µg Cat No. 611964

View Details

562077 Rev. 1

Antibody Details

47/Stat5(pY694)

Stat (Signal transducer and activators of transcription) proteins mediate the biological activity of cytokines, including interleukins, interferons, erythropoietin, and growth factors. Ligand-receptor interaction activates constitutively-associated JAK family kinases and subsequent recruitment/activation of Stat proteins by tyrosine phosphorylation. Active Stat proteins then move to the nucleus to promote transcription of cytokine-inducible genes. Seven Stat proteins have been cloned, each of which is differentially expressed and/or activated in a cytokine-specific and cell type-specific manner. Stat5 has been characterized and shown to be encoded by two separate genes, Stat5a and Stat5b, which share over 90% identity at the amino acid level. Stat5a has been shown to be involved in lactogenesis and mammary development, while Stat5b has been shown to be involved in growth hormone signaling and liver gene expression. Both Stat5a and Stat5b are involved in IL-2 induced peripheral T cell proliferation. The peptide hormone, prolactin, binds to the prolactin receptor (PRLR) to initiate the lactogenic response. There are at least three forms of PRLR; however, only the long form activates the 92-kDa Stat5 protein by inducing phosphorylation at Y694. Once phosphorylated, Stat5 becomes an essential transcription factor which binds to the β-casein gene promoter. The presence of an SH2 domain within Stat5 suggests that it may directly interact with protein tyrosine kinases (PTKs) such as JAK2.

The 47 monoclonal antibody recognizes the phosphorylated Y694 of Stat5a. The homologous phosphorylation site in Stat5b is Y699.

562077 Rev. 1

Format Details

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.

Format: PE

Excitation Source: Yellow-Green 488 nm, 532 nm, 561 nm

Excitation Max: 496 nm, 566 nm

Emission Max: 576 nm

562077 Rev.1

Citations & References

View product citations for antibody "562077" on CiteAb

Development References (5)

Bromberg J, Darnell JE. The role of STATs in transcriptional control and their impact on cellular function. Oncogene. 2000; 19(21):2468-2473. (Biology). View Reference
Gouilleux F, Wakao H, Mundt M, Groner B. Prolactin induces phosphorylation of Tyr694 of Stat5 (MGF), a prerequisite for DNA binding and induction of transcription. EMBO J. 1994; 13(18):4361-4369. (Biology). View Reference
Liu KD, Gaffen SL, Goldsmith MA. JAK/STAT signaling by cytokine receptors. Curr Opin Immunol. 1998; 10(3):271-278. (Biology). View Reference
Van De Wiele CJ, Marino JH, Murray BW, Vo SS, Whetsell ME, Teague TK. Thymocytes between the -Selection and Positive Selection Checkpoints Are Nonresponsive to IL-7 as Assessed by STAT-5 Phosphorylation. J Immunol. 2004; 172(7):4235-4244. (Biology). View Reference
Wakao H, Gouilleux F, Groner B. Mammary gland factor (MGF) is a novel member of the cytokine regulated transcription factor gene family and confers the prolactin response. EMBO J. 1994; 13(9):2182-2191. (Biology). View Reference

View All (5)

562077 Rev. 1

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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.