- 71-2500 - Invitrogen Antibodies STAT5B antibody

Submitted references Identification of STAT3 and STAT5 proteins in the rat suprachiasmatic nucleus and the Day/Night difference in astrocytic STAT3 phosphorylation in response to lipopolysaccharide.

Moravcová S, Červená K, Pačesová D, Bendová Z
Journal of neuroscience research 2016 Jan;94(1):99-108

STAT5 outcompetes STAT3 to regulate the expression of the oncogenic transcriptional modulator BCL6.

Walker SR, Nelson EA, Yeh JE, Pinello L, Yuan GC, Frank DA
Molecular and cellular biology 2013 Aug;33(15):2879-90

The KRAB zinc finger protein RSL1 regulates sex- and tissue-specific promoter methylation and dynamic hormone-responsive chromatin configuration.

Krebs CJ, Schultz DC, Robins DM
Molecular and cellular biology 2012 Sep;32(18):3732-42

Negative cross talk between NFAT1 and Stat5 signaling in breast cancer.

Zheng J, Fang F, Zeng X, Medler TR, Fiorillo AA, Clevenger CV
Molecular endocrinology (Baltimore, Md.) 2011 Dec;25(12):2054-64

Granulocyte/macrophage-colony-stimulating factor autoantibodies and myeloid cell immune functions in healthy subjects.

Uchida K, Nakata K, Suzuki T, Luisetti M, Watanabe M, Koch DE, Stevens CA, Beck DC, Denson LA, Carey BC, Keicho N, Krischer JP, Yamada Y, Trapnell BC
Blood 2009 Mar 12;113(11):2547-56

STAT5 represses BCL6 expression by binding to a regulatory region frequently mutated in lymphomas.

Walker SR, Nelson EA, Frank DA
Oncogene 2007 Jan 11;26(2):224-33

STAT5-mediated signals sustain a TCR-initiated gene expression program toward differentiation of CD8 T cell effectors.

Verdeil G, Puthier D, Nguyen C, Schmitt-Verhulst AM, Auphan-Anezin N
Journal of immunology (Baltimore, Md. : 1950) 2006 Apr 15;176(8):4834-42

Hypothalamic STAT proteins: regulation of somatostatin neurones by growth hormone via STAT5b.

Bennett E, McGuinness L, Gevers EF, Thomas GB, Robinson IC, Davey HW, Luckman SM
Journal of neuroendocrinology 2005 Mar;17(3):186-94

IL-5 and granulocyte-macrophage colony-stimulating factor activate STAT3 and STAT5 and promote Pim-1 and cyclin D3 protein expression in human eosinophils.

Stout BA, Bates ME, Liu LY, Farrington NN, Bertics PJ
Journal of immunology (Baltimore, Md. : 1950) 2004 Nov 15;173(10):6409-17

Stat5a is tyrosine phosphorylated and nuclear localized in a high proportion of human breast cancers.

Cotarla I, Ren S, Zhang Y, Gehan E, Singh B, Furth PA
International journal of cancer 2004 Feb 20;108(5):665-71

Isolation of unique STAT5 targets by chromatin immunoprecipitation-based gene identification.

Nelson EA, Walker SR, Alvarez JV, Frank DA
The Journal of biological chemistry 2004 Dec 24;279(52):54724-30

Granulocyte-macrophage colony-stimulating factor (GM-CSF)-induced STAT5 activation and target-gene expression during human monocyte/macrophage differentiation.

Lehtonen A, Matikainen S, Miettinen M, Julkunen I
Journal of leukocyte biology 2002 Mar;71(3):511-9

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Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of STAT5b expression using Rb x STAT5b.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Western blot analysis of STAT5B was performed by loading 20 µg of HeLa (lane1), HEK-293 (lane2) and A431 (lane3) cell lysates using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002), Novex® Sharp Pre-Stained Protein Standard (LC5800). Proteins were transferred to a PVDF membrane and blocked with 5 % skim milk for 1 hour at room temperature. STAT5 beta was detected at ~90 kDa using STAT5B Rabbit Polyclonal Antibody (Product # 71-2500) at 1 µg - 3 µg/mL in 2.5 % skim milk at 4°C overnight on a rocking platform. Goat Anti-Rabbit IgG - HRP Secondary Antibody (G21234) at 1:5000 dilution was used and chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Knockout of STAT5B was achieved by CRISPR-Cas9 genome editing using LentiArray™ Lentiviral sgRNA (Product # A32042) (Assay ID CRISPR773180_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Western blot analysis of STAT5B was performed by loading 30 µg of HeLa wild type (Lane 1), HeLa CAS9 (Lane 2), HeLa STAT5B KO (Lane 3) modified whole cell extracts (1%SDS). The blot was probed with Anti-STAT5 beta Polyclonal Antibody (Product # 71-2500) using 1 µg/mL dilution and Goat anti-Rabbit IgG (H+L), Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036). Loss of signal upon CRISPR mediated knockout (KO) using the LentiArray™ CRISPR product line confirms that antibody is specific to STAT5B.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Immunohistochemistry analysis of STAT5B showing staining in the cytoplasm and nucleus of paraffin-embedded mouse spleen tissue (right) compared to a negative control without primary antibody (left). To expose target proteins, antigen retrieval was performed using 10mM sodium citrate (pH 6.0), microwaved for 8-15 min. Following antigen retrieval, tissues were blocked in 3% H2O2-methanol for 15 min at room temperature, washed with ddH2O and PBS, and then probed with a STAT5B Rabbit Polyclonal Antibody (Product # 71-2500) diluted in 3% BSA-PBS at a dilution of 1:20 overnight at 4°C in a humidified chamber. Tissues were washed extensively in PBST and detection was performed using an HRP-conjugated secondary antibody followed by colorimetric detection using a DAB kit. Tissues were counterstained with hematoxylin and dehydrated with ethanol and xylene to prep for mounting.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: Flow cytometry analysis of STAT5B was done on HeLa cells treated with IFN gamma (150 ng/ml, 15 minutes). Cells were fixed with 70% ethanol for 10 minutes, permeabilized with 0.25% Triton™ X-100 for 20 minutes, and blocked with 5% BSA for 1 hour at room temperature. Cells were labeled with STAT5B Rabbit Polyclonal Antibody (712500, red histogram) or with rabbit isotype control (pink histogram) at 2 µg - 4 µg/million cells in 2.5% BSA. After incubation at room temperature for 2 - 3 hours, the cells were labeled with Alexa Fluor® 488 Goat Anti-Rabbit Secondary Antibody (A11008) at a dilution of 1:400 for 30 minutes at room temperature. The representative 10,000 cells were acquired and analyzed for each sample using an Attune® Acoustic Focusing Cytometer. The purple histogram represents unstained control cells and the green histogram represents no-primary-antibody control.

Supportive validation

Submitted by: Invitrogen Antibodies (provider)
Main image
Experimental details: ChIP- qPCR analysis of STAT5B was performed with 3 µg/mL of the STAT5B Rabbit Polyclonal Antibody (Product # 71-2500) on sheared chromatin from 2 million HeLa cells treated with IFN-γ (50 ng/mL) for 1h using the MAGnify™ Chromatin Immunoprecipitation System (Product # 49-2024). Normal rabbit IgG (3µg/mL) was used as a negative IP control. The purified DNA from each ChIP sample was analyzed by StepOnePlus™ Real-Time PCR System (Product # 4376600) with primers for the promoter of active AP-1 and COX2 gene, used as positive control targets, and the GAPDH gene, used as negative control target. Data is presented as fold enrichment of the antibody signal versus the negative control IgG using the comparative CT method.