Anti-Histone H4 antibody - ChIP Grade (ab10158)

Overview

  • Product nameAnti-Histone H4 antibody - ChIP Grade
    See all Histone H4 primary antibodies
  • Description
    Rabbit polyclonal to Histone H4 - ChIP Grade
  • Tested applicationsSuitable for: IP, ChIP, IHC-P, ICC/IF, WBmore details
  • Species reactivity
    Reacts with: Mouse, Rat, Human, Saccharomyces cerevisiae, Xenopus laevis, Drosophila melanogaster
    Predicted to work with: Chicken, Cow, Pig, Caenorhabditis elegans
  • Immunogen

    Synthetic peptide corresponding to Human Histone H4 aa 50 to the C-terminus (C terminal).
    Database link: P62805
    (Peptide available as ab13843)

  • Positive control
    • This antibody gave a positive control in the following lysates: Histone prep HeLa Histone This antibody gave a positive control in the following mouse lysates: NIH 3T3 (Mouse embryonic fibroblast cell line) Whole Cell MEF1 (Mouse embryonic fibroblast cell line) Whole Cell This antibody gave a positive control in the following rat lysate: PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell

Properties

Applications

Our Abpromise guarantee covers the use of ab10158 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
IP Use at an assay dependent concentration.
ChIP Use 2 µg for 25 µg of chromatin.
IHC-P Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval before commencing with IHC staining protocol.
ICC/IF 1/200.
WB Use a concentration of 1 µg/ml. Detects a band of approximately 14 kDa (predicted molecular weight: 11 kDa).Can be blocked with Human Histone H4 peptide (ab13843).

Target

Anti-Histone H4 antibody - ChIP Grade images



  • Performed under reducing conditions.

    Predicted band size : 11 kDa
  • Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25µg of chromatin, 2µg of ab10158 (blue), and 20µl of protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach for active and inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.

  • All lanes : Anti-Histone H4 antibody - ChIP Grade (ab10158) at 1 µg/ml

    Lane 1 : NIH 3T3 whole cell lysate (ab7179)
    Lane 2 : MEF1 (Mouse embryonic fibroblast cell line) Whole Cell Lysate
    Lane 3 : PC12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate

    Lysates/proteins at 10 µg per lane.

    Secondary
    IRDye 680 Conjugated Goat Anti-Rabbit IgG (H+L) at 1/10000 dilution

    Performed under reducing conditions.

    Predicted band size : 11 kDa
    Observed band size : 14 kDa (why is the actual band size different from the predicted?)
  • IHC image of Histone H4 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 ab10158, 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.
  • ab10158 staining Histone H4 in Mouse thymocytes by ICC/IF (Immunocytochemistry/immunofluorescence). Cells were fixed with formaldehyde, permeabilized with 0.1% Triton X-100 and blocked with 2% BSA for 1 hour at 25°C. Samples were incubated with primary antibody (1/150 in PBS) for 2.5 hours at 25°C. An Oregon Green®488-conjugated Goat anti-rabbit IgG polyclonal (1/200) was used as the secondary antibody.

    See Abreview

  • Histone H4 was immunoprecipitated using 0.5mg NIH3T3 whole cell lysate, 5µg of Rabbit polyclonal to Histone H4 and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
    The antibody was incubated under agitation with Protein G beads for 10min, NIH3T3 whole cell lysate lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
    Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab10158.
    Secondary: Clean blot (HRP conjugate) at 1/1000 dilution.
    Band: 14kDa: Histone H4; non specific - 52 and 85kDa: We are unsure as to the identity of this extra band.

References for Anti-Histone H4 antibody - ChIP Grade (ab10158)

This product has been referenced in:
  • Quarato G  et al. Sequential Engagement of Distinct MLKL Phosphatidylinositol-Binding Sites Executes Necroptosis. Mol Cell N/A:N/A (2016). WB ; Mouse . Read more (PubMed: 26853145) »
  • Zheng YC  et al. 1,2,3-Triazole-Dithiocarbamate Hybrids, a Group of Novel Cell Active SIRT1 Inhibitors. Cell Physiol Biochem 38:185-93 (2016). WB ; Human . Read more (PubMed: 26784898) »

See all 50 Publications for this product

Product Wall

Application Western blot
Sample Human Recombinant protein (Recombinant octamers & purified K-562 cell histone)
Gel Running Conditions Reduced Denaturing (10-20% BioRad TGX gel)
Loading amount 0.5 µg
Specification Recombinant octamers & purified K-562 cell histone
Blocking step Odyssey Blocking solution 1:1 (vol/vol) PBS as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 50% · Temperature: 25°C
Username

Dr. Ragnhild Eskeland

Verified customer

Submitted May 20 2015

Please try the below given antibodies. These are excellent products and are guaranteed against their respective targets.

http://www.abcam.com/histone-h4-antibody-chip-grade-ab10158.html

http://www.abcam.com/histone-h3-antibody-chip-gr...

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Xenopus laevis Cell lysate - whole cell (Erythrocytes)
Loading amount 50000 cells
Specification Erythrocytes
Gel Running Conditions Reduced Denaturing (12%)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 22°C
Username

Abcam user community

Verified customer

Submitted Apr 05 2013

The antibody ab10158 should be capable of detecting acetylated H4. It is raised against an amino acid sequence that has one acetylation site, K91. The antibody is a polyclonal antibody, though, so we expect it binds to multiple epitopes, even if k91 is...

Read More
Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Mouse Cell lysate - whole cell (liver)
Loading amount 50 µg
Specification liver
Gel Running Conditions Non-reduced Denaturing (12)
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 3% · Temperature: rt°C
Username

Abcam user community

Verified customer

Submitted Aug 03 2012

Thank you for your phone call.

I have checked the sequence homology for the wasp sequence with the human sequence for both Histone H3 and H4 and it is 96% and 99%, respectively.

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Western blot
Sample Human Cell lysate - other (Histones extracted from HeLa cells by acid extract)
Loading amount 10 µg
Specification Histones extracted from HeLa cells by acid extract
Gel Running Conditions Reduced Denaturing (15 % PAGE gel)
Blocking step Milk as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 5% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted May 31 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Mouse Cell (Mouse thymocytes)
Specification Mouse thymocytes
Fixative Formaldehyde
Permeabilization Yes - 0.1 % TritonX-100 for 5 minutes
Blocking step BSA as blocking agent for 1 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 25°C
Username

Abcam user community

Verified customer

Submitted May 28 2012

Abcam guarantees this product to work in the species/application used in this Abreview.
Application Immunocytochemistry/ Immunofluorescence
Sample Mouse Cell (neuronal)
Specification neuronal
Fixative Paraformaldehyde
Permeabilization Yes - 0.2% Triton x-100
Blocking step Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: rt°C
Username

Abcam user community

Verified customer

Submitted Mar 21 2012

Application Western blot
Sample Mouse Cell lysate - nuclear (Mouse embryonic fibroblasts)
Loading amount 40 µg
Specification Mouse embryonic fibroblasts
Gel Running Conditions Reduced Denaturing (4-20% Tris-Gly gel)
Blocking step LI-COR® Odyssey® Blocking Buffer as blocking agent for 45 minute(s) · Concentration: 50% · Temperature: RT°C
Username

Abcam user community

Verified customer

Submitted Feb 10 2012

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