Revision 1
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

REACTIVITY:

H M R

SENSITIVITY:

Endogenous

MW (kDa):

48

Source/Isotype:

Rabbit IgG

UniProt ID:

#P05412

Entrez-Gene Id:

3725

Product Information

Product Usage Information

For optimal ChIP 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.

Application Dilution
Western Blotting 1:1000
Immunoprecipitation 1:50
Immunofluorescence (Immunocytochemistry) 1:800 - 1:3200
Flow Cytometry (Fixed/Permeabilized) 1:100 - 1:200
Chromatin IP 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.

Specificity / Sensitivity

Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb recognizes endogenous levels of c-Jun protein only when phosphorylated at Ser63.

Species Reactivity:

Human, Mouse, Rat

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phospho-peptide corresponding to residues surrounding Ser63 of human c-Jun protein.

Background

c-Jun is a member of the Jun family containing c-Jun, JunB, and JunD, and is a component of the transcription factor activator protein-1 (AP-1). AP-1 is composed of dimers of Fos, Jun, and ATF family members and binds to and activates transcription at TRE/AP-1 elements (reviewed in 1). Extracellular signals, including growth factors, chemokines, and stress, activate AP-1-dependent transcription. The transcriptional activity of c-Jun is regulated by phosphorylation at Ser63 and Ser73 through SAPK/JNK (reviewed in 2). Knockout studies in mice have shown that c-Jun is essential for embryogenesis (3), and subsequent studies have demonstrated roles for c-Jun in various tissues and developmental processes, including axon regeneration (4), liver regeneration (5), and T cell development (6). AP-1 regulated genes exert diverse biological functions, including cell proliferation, differentiation, and apoptosis, as well as transformation, invasion and metastasis, depending on cell type and context (7-9). Other target genes regulate survival, as well as hypoxia and angiogenesis (8,10). Research studies have implicated c-Jun as a promising therapeutic target for cancer, vascular remodeling, acute inflammation, and rheumatoid arthritis (11,12).

  1. Jochum, W. et al. (2001) Oncogene 20, 2401-12.
  2. Davis, R.J. (2000) Cell 103, 239-52.
  3. Hilberg, F. et al. (1993) Nature 365, 179-81.
  4. Raivich, G. et al. (2004) Neuron 43, 57-67.
  5. Behrens, A. et al. (2002) EMBO J 21, 1782-90.
  6. Riera-Sans, L. and Behrens, A. (2007) J Immunol 178, 5690-700.
  7. Leppä, S. and Bohmann, D. (1999) Oncogene 18, 6158-62.
  8. Shaulian, E. and Karin, M. (2002) Nat Cell Biol 4, E131-6.
  9. Weiss, C. and Bohmann, D. (2004) Cell Cycle 3, 111-3.
  10. Karamouzis, M.V. et al. (2007) Mol Cancer Res 5, 109-20.
  11. Kim, S. and Iwao, H. (2003) J Pharmacol Sci 91, 177-81.
  12. Dass, C.R. and Choong, P.F. (2008) Pharmazie 63, 411-4.

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 BSA, 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

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.

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Revision 1
#91952

Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb

Western Blotting Image 1: Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from HeLa cells, untreated (-) or treated with UV (100 mJ/cm2, 2 hr recovery; +) and calf intestinal phosphatase and Lambda phosphatase (CIP/λ phosphatase, +) as indicated using Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb (upper), c-Jun (60A8) Rabbit mAb #9165 (middle), or β-Actin (D6A8) Rabbit mAb #8457 (lower).
Western Blotting Image 2: Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb Expand Image
Western blot analysis of extracts from ACHN cells, untreated (-) or UV-treated (100 mJ/cm2, indicated recovery times; +), using Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb (upper), c-Jun (60A8) Rabbit mAb #9165 (middle), or β-Actin (D6A8) Rabbit mAb #8457 (lower).
Immunoprecipitation Image 1: Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb Expand Image
Immunoprecipitation of Phospho-c-Jun (Ser63) protein from HeLa cell extracts treated with UV (100 mJ/cm2, 2 hr recovery). Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb. Western blot analysis was performed using Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb. Mouse Anti-rabbit IgG (Conformation Specific) (L27A9) mAb (HRP Conjugate) #5127 was used as a secondary antibody.
Immunofluorescence Image 1: Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb Expand Image
Confocal immunofluorescent analysis of ACHN cells, untreated (left), treated with UV (100 mJ/cm2, 2 hr recovery; center) or treated with UV and post-processed with λ-phosphatase (right), using Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb (green). Actin filaments were labeled with DyLight 650 Phalloidin #12956 (red). Samples were mounted in ProLong® Gold Antifade Reagent with DAPI #8961 (blue).
Flow Cytometry Image 1: Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb Expand Image
Flow cytometric analysis of ACHN cells, untreated (blue) or treated with UV (100 mJ/cm2, 1hr; green) using Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb (solid lines) or concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (dashed lines). Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
Chromatin Immunoprecipitation Image 1: Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb Expand Image
Chromatin immunoprecipitations were performed with cross-linked chromatin from PC-12 cells starved overnight and treated with Human β-Nerve Growth Factor (hβ-NGF) #5221 (50 ng/ml, 2 hr) and either Phospho-c-Jun (Ser63) (E6I7P) XP® Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Plus Enzymatic Chromatin IP Kit (Magnetic Beads) #9005. The enriched DNA was quantified by real-time PCR using SimpleChIP® Rat CCRN4L Promoter Primers #7983, rat DCLK1 promoter, SimpleChIP® Rat GAPDH Promoter Primers #7964. 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.