Product nameAnti-Histone H2A (phospho S1) + Histone H4 (phospho S1) antibody
DescriptionRabbit polyclonal to Histone H2A (phospho S1) + Histone H4 (phospho S1)
SpecificityBy peptide array ab14723 has been shown to detect both Histone H2A (phospho S1) and Histone H4 (phospho S1) peptides and not the equivalent unmodified peptides (please see peptide array image on this datasheet). However, by western blot, this antibody shows a preference for Histone H4 (phospho S1) as seen in the western blot images on this datasheet. If you have further questions about the specificity of ab14723, please contact our technical support team, who will be happy to help.
Tested applicationsSuitable for: WB, IP, ICC/IF, IHC-P, PepArrmore details
Species reactivityReacts with: Human, Saccharomyces cerevisiae
Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
Storage bufferpH: 7.40
Preservative: 0.02% Sodium azide
Constituents: PBS, 1% BSA
Concentration information loading...
PurityImmunogen affinity purified
Immunizing Peptide (Blocking)
Our Abpromise guarantee covers the use of ab14723 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||1/1000. Detects a band of approximately 15 kDa (predicted molecular weight: 11 kDa).|
|IP||1/50. See Abreview.|
|ICC/IF||Use at an assay dependent concentration.|
|IHC-P||Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.|
|PepArr||Use a concentration of 0.1 - 0.01 µg/ml.|
Cellular localizationHistone H2A: Nucleus. Chromosome. Histone H4: Nucleus. Chromosome.
All lanes : Anti-Histone H2A (phospho S1) + Histone H4 (phospho S1) antibody (ab14723) at 1 µg/ml
Lane 1 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 10 µg
Lane 2 : HeLa (Human epithelial carcinoma cell line) Nuclear Lysate at 10 µg
Lane 3 : HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate - Colcemid Treated at 2.5 µg
Lane 4 : Calf Thymus Histone Preparation Nuclear Lysate at 0.5 µg
Lane 5 : NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate at 10 µg
All lanes : Peroxidase AffiniPure Goat Anti-Rabbit IgG (H+L) (ab201489) at 1/50000 dilution
Developed using the ECL technique.
Performed under reducing conditions.
Predicted band size: 11 kDa
Observed band size: 14 kDa why is the actual band size different from the predicted?
Exposure time: 90 seconds
This blot was produced using a 4-12% Bis-tris gel under the MES buffer system. The gel was run at 200V for 35 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab14723 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406.
Peptide Array analysis of ab14723 at 1:5000 dilution against 40.0 – 0.6 µM peptides: Histone H2A unmodified peptide (Lane 1), Histone H2A (phospho S1) peptide (Lane 2), Histone H4 unmodified peptide (Lane 3) and Histone H4 (phospho S1) peptide (Lane 4). Results show strong binding to the phosphorylated, but not the unmodified versions of these peptides.
All lanes : Anti-Histone H2A (phospho S1) + Histone H4 (phospho S1) antibody (ab14723) at 1/1000 dilution
Lane 1 : YDH8 cka 2-8 ts grown at 30 degrees C
Lane 2 : YDH8 cka 2-8 ts grown at 38 degrees C
All lanes : HRP conjugated anti-Rabbit
Predicted band size: 11 kDa
Observed band size: 15 kDa why is the actual band size different from the predicted?
Additional bands at: 48 kDa. We are unsure as to the identity of these extra bands.
Cell lysates were prepared from a Casein Kinase 2 temperature sensitive Saccharomyces cerevisiae strain (YDH8 cka2-8ts) grown at either 30°C or 38°C. The kinase is active at °C and inactive at 38°C. Lysates were prepared from asynchronous log-phase cultures and 300µl run on a gel. The Western blot was blocked in milk for 20 minutes and incubated with a 1/1000 dilution of ab14723 for 16 hours. ab14723 detected a band of 15kDa representing Histone H4 (phospho S1) in the lysates grown at 30°C.
IHC image of Histone H2A (phospho S1) + Histone H4 (phospho S1) staining in human breast carcinoma FFPE section, performed on a BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab14723, 1µg/ml, for 8 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
This product has been referenced in:
- Ju J et al. NatD promotes lung cancer progression by preventing histone H4 serine phosphorylation to activate Slug expression. Nat Commun 8:928 (2017). Read more (PubMed: 29030587) »
- Zhang ZH et al. Dynamics of histone H2A, H4 and HS1ph during spermatogenesis with a focus on chromatin condensation and maturity of spermatozoa. Sci Rep 6:25089 (2016). WB . Read more (PubMed: 27121047) »