Anti-Histone H2A (phospho S1) + Histone H4 (phospho S1) antibody (ab14723)

Overview

  • Product nameAnti-Histone H2A (phospho S1) + Histone H4 (phospho S1) antibody
  • Description
    Rabbit 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 reactivity
    Reacts with: Human, Saccharomyces cerevisiae
  • Immunogen

    Synthetic peptide corresponding to Histone H4 aa 1-100 (phospho S1) conjugated to keyhole limpet haemocyanin. Also UniProt ID: P04908, Q93077, Q99878 (Histone H2A)
    Database link: P62805
    (Peptide available as ab14724)

Properties

  • FormLiquid
  • 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
  • Purification notesThis antibody was affinity purified using the phospho peptide ab14724, it was then cross-affinity purified using the non-phospho peptide ab14725 in order to remove reactivity with the non-phosphorylated epitope.
  • ClonalityPolyclonal
  • IsotypeIgG
  • Research areas

Associated products

Applications

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.

Application Abreviews Notes
WB 1/1000. Detects a band of approximately 15 kDa (predicted molecular weight: 11 kDa).

This antibody also gave a positive result in ELISA against the immunising peptide (ab14724). It gave a negative result in ELISA against the non-modified equivalent peptide (ab14725).

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.

Target

Anti-Histone H2A (phospho S1) + Histone H4 (phospho S1) antibody images

  • 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 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

    Secondary
    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.

  • 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

    Secondary
    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.

    This image was submitted as part of a review by Meredith Calvert

    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.

References for Anti-Histone H2A (phospho S1) + Histone H4 (phospho S1) antibody (ab14723)

This product has been referenced in:
  • Tjeertes JV  et al. Screen for DNA-damage-responsive histone modifications identifies H3K9Ac and H3K56Ac in human cells. EMBO J 28:1878-89 (2009). WB ; Human . Read more (PubMed: 19407812) »

See 1 Publication for this product

Product Wall

Abcam guarantees this product to work in the species/application used in this Abreview.
Application ChIP
Sample Saccharomyces cerevisiae Cell lysate - nuclear (S288c)
Specification S288c
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 15 minute(s) and 0 second(s)
Specification of the cross-linking agent: 1% Fromaldehyde
Detection step Real-time PCR
Username

Mr. Harsh Ja

Verified customer

Submitted Aug 02 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Saccharomyces cerevisiae Cell lysate - whole cell (S288c)
Loading amount 25 µg
Specification S288c
Gel Running Conditions Reduced Denaturing (15% gel)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: RT°C
Username

Mr. Harsh Ja

Verified customer

Submitted Aug 02 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunoprecipitation
Sample Saccharomyces cerevisiae Cell lysate - nuclear (BY4147 background)
Total protein in input 15 µg
Specification BY4147 background
Treatment 1% MMS (DNA damaging agent)
Immuno-precipitation step Protein A/G
Username

Abcam user community

Verified customer

Submitted Jul 26 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Saccharomyces cerevisiae Cell (BY4147 background)
Specification BY4147 background
Fixative Paraformaldehyde
Permeabilization Yes - 0.25% Triton X-100 in PBS
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 1% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted Jun 01 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application ChIP on chip
Sample Saccharomyces cerevisiae Cell lysate - nuclear (W303 background)
Specification W303 background
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 20 minute(s) and 0 second(s)
Specification of the cross-linking agent: 1% Formaldehyde
Detection step Microarray
Username

Abcam user community

Verified customer

Submitted May 31 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application ChIP
Sample Saccharomyces cerevisiae Cell lysate - nuclear (Yeast chromatin lysate)
Specification Yeast chromatin lysate
Type Cross-linking (X-ChIP)
Duration of cross-linking step: 20 minute(s) and 0 second(s)
Specification of the cross-linking agent: 1% Formaldehyde
Detection step Semiquantitative PCR
Username

Abcam user community

Verified customer

Submitted May 31 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Saccharomyces cerevisiae Cell lysate - nuclear (Yeast whole cell extract)
Loading amount 10 µg
Specification Yeast whole cell extract
Gel Running Conditions Reduced Denaturing (15% gel)
Blocking step Milk as blocking agent for 18 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: 4°C
Username

Abcam user community

Verified customer

Submitted May 26 2011

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Saccharomyces cerevisiae Cell lysate - whole cell (Asynchronous log-phase culture)
Specification Asynchronous log-phase culture
Blocking step Milk as blocking agent for 20 minute(s) · Concentration: 5%
Username

Dr. Meredith Calvert

Verified customer

Submitted Feb 23 2006

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"