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Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to Ser32 of human c-Fos protein.
Product Usage Information
For optimal ChIP and ChIP-seq, 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 |
| Fluorescent Western | 1:1000 |
| Immunoprecipitation | 1:100 |
| Immunofluorescence (Frozen) | 1:100 - 1:400 |
| Immunofluorescence (Immunocytochemistry) | 1:100 - 1:400 |
| Flow Cytometry (Fixed/Permeabilized) | 1:200 - 1:800 |
| Chromatin IP | 1:50 |
| Chromatin IP-seq | 1:50 |
Specificity/Sensitivity
Species Reactivity:
Human, Mouse, Rat
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 #99325.
参考图片
Chromatin immunoprecipitations were performed with cross-linked chromatin from PC-12 cells starved overnight and treated with Human β-Nerve Gowth Factor (hβ-NGF) #5221 (50ng/ml) for 2h and either Phospho-c-Fos (Ser32) (D82C12) XP® 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® Rat CCRN4L Promoter Primers #7983, rat DCLK1 promoter primers, and 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.
Western blot analysis of extracts from HeLa cells, serum-starved overnight and then either untreated or stimulated for 4 hours with TPA (12-O-Tetradecanoylphorbol-13-Acetate) #4174, using Phospho-c-Fos (Ser32) (D82C12) XP® Rabbit mAb (upper) and c-Fos (9F6) Rabbit mAb #2250 (lower). Antibody phospho-specificity is shown by treating lysates with λ-phosphatase.
Confocal immunofluorescent analysis of mouse cerebral cortex both untreated (left) and lambda-phosphatase treated (right) using Phospho-c-Fos (Ser32) (D82C12) XP® Rabbit mAb #5348 (green) and Neurofilament-L (DA2) Mouse mAb #2835 (red). Samples were mounted in ProLong® Gold Antifade Reagent with DAPI #8961 (blue).
Confocal immunofluorescent analysis of HeLa cells, serum-starved (left), TPA-treated (middle) or treated with TPA and λ-phosphatase (right), using Phospho-c-Fos (Ser32) (D82C12) XP® Rabbit mAb (green). Actin filaments have been labeled with DY-554 phalloidin (red).
Flow cytometric analysis of HeLa cells serum starved overnight, untreated (blue) or treated with TPA #4174 (200 nM, 4 hr; green) using Phospho-c-Fos (Ser32) (D82C12) 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 immunoprecipitations were performed with cross-linked chromatin from PC-12 cells starved overnight and treated with Human β-Nerve Gowth Factor (hβ-NGF) #5221 (50ng/ml) for 2h and Phospho-c-Fos (Ser32) (D82C12) XP® 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 Fbxl19 gene. For additional ChIP-seq tracks, please download the product datasheet.
Chromatin immunoprecipitations were performed with cross-linked chromatin from PC-12 cells starved overnight and treated with Human β-Nerve Gowth Factor (hβ-NGF) #5221 (50ng/ml) for 2h and Phospho-c-Fos (Ser32) (D82C12) XP® 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 1 (upper), including Fbxl19 (lower).