STAT5b Antibody [Unconjugated]

Western blot shows lysates of HeLa human cervical epithelial carcinoma cell line, Daudi human Burkitt's lymphoma cell line, NIH‑3T3 mouse embryonic fibroblast cell line, M1 mouse myeloid leukemia cell line, and PC‑12 rat adrenal pheochromocytoma cell line. PVDF membrane was probed with 0.2 µg/mL of Rabbit Anti-Human/Mouse STAT5b Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1584) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). A specific band was detected for STAT5b at approximately 90 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

STAT5b was detected in immersion fixed HepG2 human hepatocellular carcinoma cell line using Rabbit Anti-Human/Mouse STAT5b Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1584) at 5 µg/mL for 3 hours at room temperature. Cells were stained using the NorthernLights™ 557-conjugated Anti-Rabbit IgG Secondary Antibody (red; Catalog # NL004) and counterstained with DAPI(blue). Specific staining was localized to cytoplasm and nucleus. View our protocol for Fluorescent ICC Staining of Cells on Coverslips.

HeLa human cervical epithelial carcinoma and M1 mouse myeloid leukemia cell line were untreated (-) or activated (+) with 100 ng/mL Recombinant Human IFN‑ gamma (Catalog # 285-IF) for 15 minutes. STAT5b was immunoprecipitated from lysates of 5 x 106 cells following incubation with 2 µg Rabbit Anti-Human/Mouse STAT5b Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1584) for 1 hour at room temperature. STAT5b-antibody complexes were absorbed using Protein A sepharose (Invitrogen, Catalog # 10-1041). Immunoprecipitated STAT5b was detected by Western blot using 1 µg/mL Human Phospho-STAT5a/b (Y699) Antigen Affinity-purified Polyclonal Antibody (Catalog # AF4190). View our recommended buffer recipes for immunoprecipitation.

Jurkat human acute T cell leukemia cell line was stained with Rabbit Anti-Human/Mouse STAT5b Antigen Affinity‑purified Polyclonal Antibody (Catalog # AF1584, filled histogram) or control antibody (Catalog # AB-105-C, open histogram), followed by Allophycocyanin-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # F0111). To facilitate intracellular staining, cells were fixed with paraformaldehyde and permeabilized with methanol.

Simple Western lane view shows lysates of HeLa human cervical epithelial carcinoma cell line and NIH‑3T3 mouse embryonic fibroblast cell line, loaded at 0.2 mg/mL. A specific band was detected for STAT5b at approximately 92 kDa (as indicated) using 10 µg/mL of Rabbit Anti-Human/Mouse STAT5b Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1584). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.Non-specific interaction with the 230 kDa Simple Western standard may be seen with this antibody.

Western blot shows lysates of HeLa human cervical epithelial carcinoma parental cell line and STAT5b knockout HeLa cell line (KO). PVDF membrane was probed with 0.2 µg/mL of Rabbit Anti-Human/Mouse STAT5b Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1584) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody (Catalog # HAF008). A specific band was detected for STAT5b at approximately 90 kDa (as indicated) in the parental HeLa cell line, but is not detectable in knockoutHeLa cell line. GAPDH (Catalog # AF5718) is shown as a loading control. This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Western blot shows lysates of HAP1 human near-haploid cell line and STAT5b knockout HAP1 cell line (KO). Nitrocellulose membrane was probed with 0.2 µg/mL of Rabbit Anti-Human/Mouse STAT5b Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1584) followed by HRP-conjugated Anti-Rabbit IgG Secondary Antibody. A specific band was detected for STAT5b at approximately 85 kDa (as indicated) in the parental HAP1 cell line, but is not detectable in knockout HAP1 cell line. The Ponceau stained transfer of the blot is shown. This experiment was conducted under reducing conditions. Image, protocol, and testing courtesy of YCharOS Inc. See ycharos.com for additional details.

Immunoprecipitation was performed on cell lysate of HAP1 human near-haploid cell line using 2.0 μg of Rabbit Anti-Human Stat5b Polyclonal Antibody (Catalog # AF1584) pre-coupled to protein G or protein A beads. Immunoprecipitated Stat5b was detected with a Mouse Anti-Stat5b antibody. The Ponceau stained transfers of each blot are shown. SM=10% starting material; UB=10% unbound fraction; IP=immunoprecipitated. Image, protocol, and testing courtesy of YCharOS Inc. (ycharos.com).

• Reactivity: Hu, Mu
• Applications: WB, Simple Western, IP, CyTOF-ready, Flow, ICC/IF, KO
• Clonality: Polyclonal
• Host: Rabbit
• Conjugate: Unconjugated
• Concentration: LYOPH

STAT5b Antibody [Unconjugated] Summary

• Immunogen: Human STAT5b synthetic peptide
  aa 777-787
• Specificity: Detects human and mouse STAT5b.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Rabbit
• Gene: STAT5B
• Purity Statement: Antigen Affinity-purified

Applications/Dilutions

• Dilutions:
  • CyTOF-ready
  • Immunocytochemistry 5-15 ug/mL
  • Immunoprecipitation 1-5 ug/500 ug cell lysate
  • Intracellular Staining by Flow Cytometry 2.5 ug/10^6 cells
  • Knockout Validated
  • Simple Western 10 ug/mL
  • Western Blot 0.2 ug/mL
• Application Notes: In Simple Western only 10-15 uL of the recommended dilution is used per data point.

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.
• Buffer: Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Reconstitute at 0.2 mg/mL in sterile PBS.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for STAT5b Antibody [Unconjugated]

• signal transducer and activator of transcription 5B
• STAT5
• STAT5b
• transcription factor STAT5B

Background

Signal transduction and activator of transcription 5 (STAT5) is a member of the Jak/STAT signal transduction pathway and is activated by a variety of cytokines (IL22, IL6, IFN-α). STAT5 has two isoforms (A and B) that share 93% amino acid identity and bind the DNA consensus site TTCN3GAA. STAT5 mediates cytokine signaling by acting as a signal transducer in the cytoplasm and, upon phosphorylation, translocates to the nucleus and activates transcription of specific genes. STAT5 is involved in a wide array of biological processes ranging from regulating apoptosis to adult mammary gland proliferation, differentiation and survival.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for STAT5b Antibody (AF1584)(3)

Publications using AF1584

• JX Lin, N Du, P Li, M Kazemian, T Gebregiorg, R Spolski, WJ Leonard Critical functions for STAT5 tetramers in the maturation and survival of natural killer cells Nat Commun, 2017-11-06;8(1):1320. 2017-11-06 [PMID: 29105654] (Immunoprecipitation, Mouse)
• Schaller-Schonitz M, Barzan D, Williamson A, Griffiths J, Dallmann I, Battmer K, Ganser A, Whetton A, Scherr M, Eder M BCR-ABL affects STAT5A and STAT5B differentially. PLoS ONE, 2014-05-16;9(5):e97243. 2014-05-16 [PMID: 24836440] (Immunoprecipitation, Mouse)
• Bao YC, Nomura Y, Moon Y, Tonozuka Y, Minoshima Y, Hatori T, Tsuchiya A, Kiyono M, Nosaka T, Nakajima H Rac1 and a GTPase-activating protein, MgcRacGAP, are required for nuclear translocation of STAT transcription factors. J. Cell Biol., 2006-12-18;175(6):937-46. 2006-12-18 [PMID: 17178910] (ICC, Western Blot, Human, Mouse)

Review for STAT5b Antibody (AF1584) (1)

Average Rating: 4

(Based on 1 review)

We have 1 review tested in 1 species: Human.

reviewed by: Verified Customer WB Human 01/05/2015

Summary

• Application: Western Blot
• Sample Tested: See PMID 22729867
• Species: Human

Bioinformatics

• Gene Symbol: STAT5B