Revision 6

#72604Store at -20C

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Applications:

WB, IP, IF-IC, FC-FP, ChIP, ChIP-seq, C&R

REACTIVITY:

H M

SENSITIVITY:

Endogenous

MW (kDa):

97, 113

Source/Isotype:

Rabbit IgG

UniProt ID:

#P52630

Entrez-Gene Id:

6773

Product Information

Product Usage Information

For optimal ChIP and ChIP-seq results, use 10 μl of antibody and 10 μg of chromatin (approximately 4 x 106 cells) per IP. This antibody has been validated using SimpleChIP® Enzymatic Chromatin IP Kits.

The CUT&RUN dilution was determined using CUT&RUN Assay Kit #86652.
Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50
Immunofluorescence (Immunocytochemistry) 1:100 - 1:400
Flow Cytometry (Fixed/Permeabilized) 1:50 - 1:200
Chromatin IP 1:50
Chromatin IP-seq 1:50
CUT&RUN 1:50

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

For a carrier free (BSA and azide free) version of this product see product #19413.

Specificity / Sensitivity

Stat2 (D9J7L) Rabbit mAb recognizes endogenous levels of total Stat2 protein.

Species Reactivity:

Human, Mouse

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu706 of human Stat2 protein.

Background

Stat2 (113 kDa), originally purified from the nuclei of alpha-interferon-treated cells, is critical to the transcriptional responses induced by type I interferons, IFN-alpha/beta (1,2). Knockout mice with a targeted disruption of Stat2 have higher susceptibility to viral infection and altered responses to type I interferons (3). Stat2 is rapidly activated by phosphorylation at Tyr690 in response to stimulation by IFN-alpha/beta via associations with receptor-bound Jak kinases (4). Unlike other Stat proteins, Stat2 does not form homodimers. Instead, activated Stat2 forms a heterodimer with Stat1 and translocates to the nucleus. There, it associates with the DNA-binding protein p48 and forms the transcriptional activator complex, interferon-stimulated gene factor 3 (ISGF3), promoting transcription from the ISRE (5).

  1. Fu, X.Y. et al. (1992) Proc Natl Acad Sci U S A 89, 7840-3.
  2. Ihle, J.N. (2001) Curr Opin Cell Biol 13, 211-7.
  3. Park, C. et al. (2000) Immunity 13, 795-804.
  4. Improta, T. et al. (1994) Proc Natl Acad Sci U S A 91, 4776-80.
  5. Horvath, C.M. et al. (1996) Mol Cell Biol 16, 6957-64.

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

WB: Western Blotting IP: Immunoprecipitation IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) ChIP: Chromatin IP ChIP-seq: Chromatin IP-seq C&R: CUT&RUN

Cross-Reactivity Key

H: human M: mouse R: rat Hm: hamster Mk: monkey Vir: virus Mi: mink C: chicken Dm: D. melanogaster X: Xenopus Z: zebrafish B: bovine Dg: dog Pg: pig Sc: S. cerevisiae Ce: C. elegans Hr: horse GP: Guinea Pig Rab: rabbit All: all species expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Revision 6
#72604

Stat2 (D9J7L) Rabbit mAb

Western Blotting Image 1: Stat2 (D9J7L) Rabbit mAb Expand Image
Western blot analysis of extracts from various cell lines using Stat2 (D9J7L) Rabbit mAb. KARPAS cell line source: Dr Abraham Karpas at the University of Cambridge.
Immunoprecipitation Image 1: Stat2 (D9J7L) Rabbit mAb Expand Image
Immunoprecipitation of Stat2 from KARPAS-299 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Stat2 (D9J7L) Rabbit mAb. Western blot was performed using Stat2 (D9J7L) Rabbit mAb. KARPAS cell line source: Dr Abraham Karpas at the University of Cambridge.
Immunofluorescence Image 1: Stat2 (D9J7L) Rabbit mAb Expand Image
Confocal immunofluorescent analysis of A-431 cells, serum starved (left) or treated with IFNα (1000 U/ml for 30 mins; right) using Stat2 (D9J7L) Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
Flow Cytometry Image 1: Stat2 (D9J7L) Rabbit mAb Expand Image
Flow cytometric analysis of K-562 cells using Stat2 (D9J7L) Rabbit mAb (solid line) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype control #3900 (dashed line). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Chromatin Immunoprecipitation Image 1: Stat2 (D9J7L) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from U266 cells treated with Human Interferon-α (IFN-α) #9906 (10nM) for 30 min and Stat2 (D9J7L) Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across USP18, a known target gene of Stat2 (see additional figure containing ChIP-qPCR data). For additional ChIP-seq tracks, please download the product datasheet.
Chromatin Immunoprecipitation Image 2: Stat2 (D9J7L) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from U266 cells treated with Human Interferon-α (IFN-α) #9906 (10nM) for 30 min and Stat2 (D9J7L) Rabbit mAb, using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. DNA Libraries were prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across chromosome 22 (upper), including USP18 (lower), a known target gene of Stat2 (see additional figure containing ChIP-qPCR data).
Chromatin Immunoprecipitation Image 3: Stat2 (D9J7L) Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from U266 cells treated with Human Interferon-α (IFN-α) #9906 (100 ng/ml) for 30 min, and either Stat2 (D9J7L) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using human USP18 promoter primers, SimpleChIP® Human WARS Intron 1 Primers #30101, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
CUT and RUN Image 1: Stat2 (D9J7L) Rabbit mAb Expand Image
CUT&RUN was performed with U266 cells treated with Human Interferon-α1 (hIFN-α1) #8927 (10nM, 30min) and Stat2 (D9J7L) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Library was prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figure shows binding across GAS5 gene.
CUT and RUN Image 2: Stat2 (D9J7L) Rabbit mAb Expand Image
CUT&RUN was performed with U266 cells treated with Human Interferon-α1 (hIFN-α1) #8927 (10nM, 30min) and Stat2 (D9J7L) Rabbit mAb, using CUT&RUN Assay Kit #86652. DNA Library was prepared using DNA Library Prep Kit for Illumina® (ChIP-seq, CUT&RUN) #56795. The figures show binding across chromosome 1 (upper), including GAS5 gene (lower).
CUT and RUN Image 3: Stat2 (D9J7L) Rabbit mAb Expand Image
CUT&RUN was performed with U266 cells treated with Human Interferon-α1 (hIFN-α1) #8927 (10nM, 30min) and either Stat2 (D9J7L) Rabbit mAb or Rabbit (DA1E) mAb IgG XP® Isotype Control (CUT&RUN) #66362, using CUT&RUN Assay Kit #86652. The enriched DNA was quantified by real-time PCR using human SIN3A promoter primers and human ITM2A upstream primers. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.