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Phospho-STAT5 (Y694) Recombinant Rabbit Monoclonal Antibody [SC05-31]

Usd: 385

Specification

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Catalog# ET1610-48

Phospho-STAT5 (Y694) Recombinant Rabbit Monoclonal Antibody [SC05-31]

Application

  • WB

  • IHC-P

  • IF-Cell

Reactivity

  • Human

  • Rat

BSA and Azide free
Predicted reactivity

  • Mouse

Predicted species support after-sales service
Conjugation

  • unconjugated

This product has been cited in peer reviewed publications, see list HERE

Overview

Product Name

Phospho-STAT5 (Y694) Recombinant Rabbit Monoclonal Antibody [SC05-31]

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Synthetic phospho-peptide corresponding to residues surrounding Tyr694 of human STAT5A.

Species Reactivity

Human, Rat (Predicted: Mouse)

Predicted species support after-sales service

Validated Applications

WB, IHC-P, IF-Cell

Molecular Weight

Predicted band size: 91 kDa

Positive Control

PC-12 treated with 1mM Sodium orthovanadate for 30 minutes cell lysate, PC-12 cells treated with 1mM Sodium orthovanadate for 30 minutes, Hela cells treated with 100ng/mL IFN alpha for 60 minutes, human breast carcinoma tissue.

Conjugation

unconjugated

Clone Number

SC05-31

RRID

AB_3069933

Product Features

Form

Liquid

Concentration

Storage Instructions

Shipped at 4℃. Store at +4℃ short term (1-2 weeks). It is recommended to aliquot into single-use upon delivery. Store at -20℃ long term.

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

  • WB

  • 1:1,000-1:2,000

  • IHC-P

  • 1:30,000

  • IF-Cell

  • 1:100

Target

Function

Stat5 (Signal Transducers and Activators of Transcription 5) is important in regulating T cell functions involving the receptors for Interleukin-2 (IL-2). IL-2 stimulates the rapid phosphorylation of both serine and tyrosine residues of Stat5a and Stat5b in human T lymphocytes and in several IL-2-responsive lymphocytic cell lines. IL-2 differentially induces serine phosphorylation of Stat5a and Stat5b on Ser726 and Ser731, respectively. Stat5b is preferentially phosphorylated and displays more protracted serine phosphorylation kinetics than Stat5a. Both the acid-rich region and the COOH terminus of IL-2Rb can independently mediate IL-2-induced Stat 5a/b serine phosphorylation, suggesting that Stat5a/b serine phosphorylation occurs at a postreceptor level. Stat5a is phosphorylated on Tyr694 in a prolactin-sensitive manner, whereas serine phosphorylation is constitutive. Activation of Stat5 by IL-2 may help govern the biological effects of IL-2 during the immune response.

Background References

1. Yang TY et al. A multiple reaction monitoring (MRM) method to detect Bcr-Abl kinase activity in CML using a peptide biosensor. PLoS One 8:e56627 (2013).

2. Caldarelli A et al. A genome-wide RNAi screen identifies proteins modulating aberrant FLT3-ITD signaling. Leukemia 27:2301-10 (2013).

Sequence Similarity

Belongs to the transcription factor STAT family.

Post-translational Modification

Tyrosine phosphorylated in response to KITLG/SCF, IL2, IL3, IL7, IL15, CSF2/GMCSF, GH1, PRL, EPO and THPO (By similarity). Activated KIT promotes phosphorylation on tyrosine residues and subsequent translocation to the nucleus. Tyrosine phosphorylated in response to constitutively activated FGFR1, FGFR2, FGFR3 and FGFR4 (By similarity). Tyrosine phosphorylation is required for DNA-binding activity and dimerization. Serine phosphorylation is also required for maximal transcriptional activity (By similarity). Tyrosine phosphorylated in response to signaling via activated FLT3; wild-type FLT3 results in much weaker phosphorylation than constitutively activated mutant FLT3. Alternatively, can be phosphorylated by JAK2 at Tyr-694.; ISGylated.

Subcellular Location

Cytoplasm, Nucleus.

UNIPROT

SwissProt: P42229 Human

SwissProt: P51692 Human

SwissProt: P42230 Mouse

SwissProt: P42232 Mouse

SwissProt: P52632 Rat

SwissProt: Q62771 Rat

Synonyms

MGF antibody

Signal transducer and activator of transcription 5A antibody

Signal Transducer and Activator of Transcription 5B antibody

STA5A_HUMAN antibody

STAT 5A antibody

STAT 5B antibody

STAT5 antibody

STAT5A antibody

STAT5B antibody

Transcription factor STAT5A antibody

Expand

Images

  • <span style="font-weight: bold;">☑ Cell treatment (CT)</span><br /><br />Western blot analysis of Phospho-STAT5 (Y694) on different lysates with Rabbit anti-Phospho-STAT5 (Y694) antibody (<a href="/products/ET1610-48" style="font-weight: bold;text-decoration: underline;">ET1610-48</a>) at 1/1,000 dilution.<br /><br />Lane 1: PC-12 cell lysate<br />Lane 2: PC-12 treated with 1mM Sodium orthovanadate for 30 minutes cell lysate<br />Lane 3: PC-12 treated with 1mM Sodium orthovanadate for 30 minutes cell lysate, then the membrane treated with λpp for 1 hour<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 91 kDa<br />Observed band size: 91 kDa<br /><br />Exposure time: 3 minutes;<br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1610-48" style="font-weight: bold;text-decoration: underline;">ET1610-48</a>) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    ☑ Cell treatment (CT)

    Western blot analysis of Phospho-STAT5 (Y694) on different lysates with Rabbit anti-Phospho-STAT5 (Y694) antibody (ET1610-48) at 1/1,000 dilution.

    Lane 1: PC-12 cell lysate
    Lane 2: PC-12 treated with 1mM Sodium orthovanadate for 30 minutes cell lysate
    Lane 3: PC-12 treated with 1mM Sodium orthovanadate for 30 minutes cell lysate, then the membrane treated with λpp for 1 hour

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 91 kDa
    Observed band size: 91 kDa

    Exposure time: 3 minutes;
    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1610-48) at 1/1,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • <span style="font-weight: bold;">☑ Cell treatment (CT)</span><br /><br />Immunocytochemistry analysis of PC-12 cells treated with 1mM Sodium orthovanadate for 30 minutes labeling Phospho-STAT5 (Y694) with Rabbit anti-Phospho-STAT5 (Y694) antibody (<a href="/products/ET1610-48" style="font-weight: bold;text-decoration: underline;">ET1610-48</a>) at 1/100 dilution.<br /><br />Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-STAT5 (Y694) antibody (<a href="/products/ET1610-48" style="font-weight: bold;text-decoration: underline;">ET1610-48</a>) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor&trade; 488, <a href="/products/HA1121" style="font-weight: bold;text-decoration: underline;">HA1121</a>) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.<br /><br />Beta tubulin (<a href="/products/M1305-2" style="font-weight: bold;text-decoration: underline;">M1305-2</a>, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor&trade; 594, <a href="/products/HA1126" style="font-weight: bold;text-decoration: underline;">HA1126</a>) was used as the secondary antibody at 1/1,000 dilution.

    ☑ Cell treatment (CT)

    Immunocytochemistry analysis of PC-12 cells treated with 1mM Sodium orthovanadate for 30 minutes labeling Phospho-STAT5 (Y694) with Rabbit anti-Phospho-STAT5 (Y694) antibody (ET1610-48) at 1/100 dilution.

    Cells were fixed in 4% paraformaldehyde for 20 minutes at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-Phospho-STAT5 (Y694) antibody (ET1610-48) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.

    Beta tubulin (M1305-2, red) was stained at 1/100 dilution overnight at +4℃. Goat Anti-Mouse IgG H&L (iFluor™ 594, HA1126) was used as the secondary antibody at 1/1,000 dilution.

  • <span style="font-weight: bold;">☑ Cell treatment (CT)</span><br /><br />Western blot analysis of Phospho-STAT5(Y694) on Hela cell lysates.<br /><br />Lane 1: Hela cells, whole cell lysate, 10ug/lane<br />Lane 2/3: Hela cells treated with 100ng/mL IFN alpha for 60 minutes, whole cell lysates, 10ug/lane<br />Lane 4: Hela cells treated with 100ng/mL IFN alpha for 60 minutes, then treated with 2.8ug/ul lambda-PP for 30 minutes, whole cell lysates, 10ug/lane<br /><br />All lanes :<br />Anti-Phospho-STAT5(Y694) antibody (<a href="/products/ET1610-48" style="font-weight: bold;text-decoration: underline;">ET1610-48</a>) at 1:500 dilution. Anti-STAT5 antibody (<a href="/products/ET1612-63" style="font-weight: bold;text-decoration: underline;">ET1612-63</a>) at 1:500 dilution. Anti-GAPDH antibody (<a href="/products/ET1601-4" style="font-weight: bold;text-decoration: underline;">ET1601-4</a>) at 1:10,000 dilution. Goat Anti-Rabbit IgG H&L (HRP) (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/200,000 dilution.<br /><br />Predicted band size: 91 kDa<br />Observed band size: 91 kDa<br /><br />Blocking and diluting buffer: 5% BSA.<br />Exposure time: 2 minutes 2 seconds

    ☑ Cell treatment (CT)

    Western blot analysis of Phospho-STAT5(Y694) on Hela cell lysates.

    Lane 1: Hela cells, whole cell lysate, 10ug/lane
    Lane 2/3: Hela cells treated with 100ng/mL IFN alpha for 60 minutes, whole cell lysates, 10ug/lane
    Lane 4: Hela cells treated with 100ng/mL IFN alpha for 60 minutes, then treated with 2.8ug/ul lambda-PP for 30 minutes, whole cell lysates, 10ug/lane

    All lanes :
    Anti-Phospho-STAT5(Y694) antibody (ET1610-48) at 1:500 dilution. Anti-STAT5 antibody (ET1612-63) at 1:500 dilution. Anti-GAPDH antibody (ET1601-4) at 1:10,000 dilution. Goat Anti-Rabbit IgG H&L (HRP) (HA1001) at 1/200,000 dilution.

    Predicted band size: 91 kDa
    Observed band size: 91 kDa

    Blocking and diluting buffer: 5% BSA.
    Exposure time: 2 minutes 2 seconds

  • Immunohistochemical analysis of paraffin-embedded human breast cancer tissue with Rabbit anti-Phospho-STAT5 (Y694) antibody (<a href="/products/ET1610-48" style="font-weight: bold;text-decoration: underline;">ET1610-48</a>) at 1/30,000 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/ET1610-48" style="font-weight: bold;text-decoration: underline;">ET1610-48</a>) at 1/30,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human breast cancer tissue with Rabbit anti-Phospho-STAT5 (Y694) antibody (ET1610-48) at 1/30,000 dilution.

    The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH 6.0) (high pressure) for 2 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (ET1610-48) at 1/30,000 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

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Citation

Products with the same target and pathway

metafield image

Phospho-STAT5 (Y694) Recombinant Rabbit Monoclonal Antibody [SC05-31] - BSA and Azide free

Application: WB,IHC-P,IF-Cell

Reactivity: Human,Rat,Mouse

Conjugate: unconjugated