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9939
Stat Antibody Sampler Kit
Primary Antibodies
Antibody Sampler Kit

Stat Antibody Sampler Kit #9939

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Stat Antibody Sampler Kit: Image 1

Chromatin immunoprecipitations were performed with cross-linked chromatin from HT-1080 cells treated with IFN-γ (50 ng/ml) for 30 minutes and either Stat1 Antibody 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 IRF-1 promoter primers, SimpleChIP® Human TAP1 Promoter Primers #5148, 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.

Stat Antibody Sampler Kit: Image 2

Western blot analysis of extracts from A549 cells (lane 1) or STAT1 knock-out cells (lane 2) using Stat1 Antibody #9172 (upper), and β-actin (D6A8) Rabbit mAb #8457 (lower). The absence of signal in the STAT1 knock-out A549 cells confirms specificity of the antibody for STAT1.

Stat Antibody Sampler Kit: Image 3

Western blot analysis of extracts from HeLa cells, transfected with 100 nM SignalSilence® Control siRNA (Unconjugated) #6568 (-), SignalSilence® Stat3 siRNA I (+) or SignalSilence® Stat3 siRNA II #6582 (+), using Stat3 (79D7) Rabbit mAb #4904 and α-Tubulin (11H10) Rabbit mAb #2125. The Stat3 (79D7) Rabbit mAb confirms silencing of Stat3 expression, while the α-Tubulin (11H10) Rabbit mAb is used to control for loading and specificity of Stat3 siRNA.

Stat Antibody Sampler Kit: Image 4

Chromatin immunoprecipitations were performed with cross-linked chromatin from Hep G2 cells treated with IL-6 (100 ng/ml) for 30 minutes, and either Stat3 (79D7) 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 IRF-1 promoter primers, SimpleChIP® Human c-Fos Promoter Primers #4663, 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.

Stat Antibody Sampler Kit: Image 5

Chromatin immunoprecipitations were performed with cross-linked chromatin from BaF3 cells starved of IL-3 for 6 hours followed by induction with IL-3 for 45 minutes, and either Stat5 (D2O6Y) 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 SimpleChIP® Mouse CIS Intron 1 Primers #5131, mouse SOCS-3 promoter primers, and SimpleChIP® Mouse RPL30 Intron 2 Primers #7015. 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.

Stat Antibody Sampler Kit: Image 6

Chromatin immunoprecipitations were performed with cross-linked chromatin from Ramos cells starved overnight then treated with IL-4 (100 ng/ml, 30 min) and either Stat6 (D3H4) 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 SimpleChIP® Human DMD Intron 2 Primers #7710, human MS4A1 promoter primers, 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.

Stat Antibody Sampler Kit: Image 7

After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.

Stat Antibody Sampler Kit: Image 8

Western blot analysis of extracts from HeLa cells 48 hours following mock transfection, transfection with non-targeted (control) siRNA or transfection with Stat1 siRNA. Stat1 was detected using Stat1 Antibody #9172, and p42 was detected using p42 MAPK Antibody #9108. The Stat1 Antibody confirms silencing of Stat1 expression, and the p42 MAPK Antibody is used to control for protein loading and siRNA specificity.

Stat Antibody Sampler Kit: Image 9

Western blot analysis of extracts from various cell lines using Stat3 (79D7) Rabbit mAb.

Stat Antibody Sampler Kit: Image 10

Immunoprecipitation of Stat5 from K-562 cell extracts. Lane 1 is 10% input, lane 2 is precipitated with Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Stat5 (D2O6Y) Rabbit mAb. Western blot was performed using Stat5 (D2O6Y) Rabbit mAb.

Stat Antibody Sampler Kit: Image 11

Western blot analysis of extracts from various cell lines using Stat6 (D3H4) Rabbit mAb.

Stat Antibody Sampler Kit: Image 12

Western blot analysis of extracts from SK-MEL-28 cells, untreated or IFN-alpha-treated (100 ng/ml), using Phospho-Stat1 (Tyr701) Antibody #9171 (upper) or Stat1 Antibody (lower).

Stat Antibody Sampler Kit: Image 13

Western blot analysis of extracts from various cell lines using Stat5 (D2O6Y) Rabbit mAb (upper) and β-Actin (D6A8) Rabbit mAb #8457 (lower).

Stat Antibody Sampler Kit: Image 14

Western blot analysis of PC-3 cells, mock transfected (-) or transfected with constructs expressing full-length human Stat5a (+) or Stat5b (+) using Stat5 (D2O6Y) Rabbit mAb.

To Purchase # 9939T
Product # Size Price
9939T
1 Kit  (4 x 20 µl) $ 327

Product Includes Quantity Applications Reactivity MW(kDa) Isotype
Stat1 Antibody 9172 20 µl
  • WB
  • IP
  • ChIP
H M R Mk 84, 91 Rabbit 
Stat3 (79D7) Rabbit mAb 4904 20 µl
  • WB
  • IP
  • ChIP
H M R Mk 79, 86 Rabbit IgG
Stat5 (D2O6Y) Rabbit mAb 94205 20 µl
  • WB
  • IP
  • ChIP
H M R 90 Rabbit IgG
Stat6 (D3H4) Rabbit mAb 5397 20 µl
  • WB
  • IP
  • ChIP
H M R 110 Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl
  • WB
Goat 

Product Description

The Stat Antibody Sampler Kit provides an economical means to examine multiple Stat proteins: Stat1, Stat3, Stat5 and Stat6. The kit contains enough primary and secondary antibodies to perform two Western blot experiments.

Specificity / Sensitivity

Each Stat antibody in the kit recognizes only its target protein, independent of phosphorylation state.

Source / Purification

Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to the sequence of human Stat1. Polyclonal antibodies are purified by protein A and peptide affinity chromatography. Monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Leu60 of human Stat5, Gly582 of human Stat6, and a Stat3 fusion protein corresponding to the carboxy-terminal sequence of mouse Stat3.

Background

Jaks (Janus Kinases) and Stats (Signal Transducers and Activators of Transcription) are utilized by receptors for a wide variety of ligands including cytokines, hormones, growth factors and neurotransmitters. Jaks, activated via autophosphorylation following ligand-induced receptor aggregation, phosphorylate tyrosine residues on associated receptors, Stat molecules and other downstream signaling proteins (1,2). The phosphorylation of Stat proteins at conserved tyrosine residues activates SH2-mediated dimerization followed rapidly by nuclear translocation. Stat dimers bind to IRE (interferon response element) and GAS (gamma interferon-activated sequence) DNA elements, resulting in the transcriptional regulation of downstream genes (1,2). The remarkable range and specificity of responses regulated by the Stats is determined in part by the tissue-specific expression of different cytokine receptors, Jaks and Stats (2,3), and by the combinatorial coupling of various Stat members to different receptors. Serine phosphorylation in the carboxy-terminal transcriptional activation domain has been shown to regulate the function of Stat1, -2, -3, -4 and -5 (1). Phosphorylation of Stat3 at Ser727 via MAPK or mTOR pathways is required for optimal transcriptional activation in response to growth factors and cytokines including IFN-gamma and CNTF (4,5). Jak/Stat pathways also play important roles in oncogenesis, tumor progression, angiogenesis, cell motility, immune responses and stem cell differentiation (6-11).

  1. Darnell, J.E. et al. (1994) Science 264, 1415-21.
  2. Leonard, W.J. and O'Shea, J.J. (1998) Annu Rev Immunol 16, 293-322.
  3. Caldenhoven, E. et al. (1996) J. Biol. Chem. 271, 13221-13227.
  4. Wen, Z. et al. (1995) Cell 82, 241-50.
  5. Yokogami, K. et al. (2000) Curr Biol 10, 47-50.
  6. Lim, C.P. and Cao, X. (1999) J. Biol. Chem. 274, 31055-31061.
  7. Bromberg, J.F. et al. (1999) Cell 98, 295-303.
  8. Su, L. et al. (1999) J. Biol. Chem. 274, 31770-31774.
  9. Dentelli, P. et al. (1999) J Immunol 163, 2151-9.
  10. Cattaneo, E. et al. (1999) Trends Neurosci. 22, 365-369.
  11. Frank, D.A. (1999) Mol Med 5, 432-56.

Limited Uses

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For Research Use Only. Not For Use In Diagnostic Procedures.
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