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Product name
Anti-Histone H4 (tri methyl K20) antibody - ChIP Grade
See all Histone H4 products (10) ...
Description
Rabbit polyclonal to Histone H4 (tri methyl K20) - ChIP Grade
Specificity
Histone H4 tri-methylated at Lys 20.
Tested applications
ChIP/Chip, IHC-P, Electron Microscopy, IHC (PFA fixed), IHC-Fr, ICC/IF, Flow Cyt, IF, WB, ChIPmore details
Cross reactivity
Reacts with
Mouse, Rat, Cow, Human, Schizosaccharomyces pombe, Toxoplasma gondii
Predicted to work with
all Mammals
Immunogen
Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H4, tri methylated at K20.
(Peptide available as ab175 67.)
Positive control
Calf Thymus Histone Preparation; Hela whole cell extract
Properties
Form
Liquid
Storage instructions
Store at +4°C short term (1-2 weeks). Aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
Storage buffer
Preservative: 0.02% Sodium Azide
Constituents: 1% BSA, PBS, pH 7.4
Concentration
Concentration information loading...
Purity
Immunogen affinity purified
Purification notes
Purified using Sulpholink column with specific peptide linked via its cysteine residue
Primary antibody notes
In immunofluorescence: Human cells: The antibody stains a subset of cellular chromatin that is characteristically DAPI-rich (condensed, heterochromatic). The perinucleolar heterochromatin is particularly rich in trimethylated lysine 20 staining. Mouse: Prominent staining of a subset of centromeric or pericentromeric heterochromatin. Interphase: The cells within the culture show a considerable variability in the intensity of staining with the antibody. The relationship between trimethylation levels and cell cycle have not yet been determined but may be a contributor to the amount of methylation detected in each cell. Heterochromatic regions of the interphase nucleus are the primary sites of trimethylation observed by indirect immunofluroescence. Mitosis: Discrete chromosomal regions are labelled intensely, with lower level fluorescence throughout the remainder of the chromosome arms. Immunofluorescence staining was performed as part of the nuclear antibody characterisation program at www.cellnucleus.com. Trimenthylation at Lys20 of Histone H4 isa common hallmark of human cancer (Fraga et al. 2005).
Clonality
Polyclonal
Isotype
IgG
Research areas
Epigenetics and Nuclear Signaling >> ChIP'ing antibodies >> ChIP'ing antibodies
Epigenetics and Nuclear Signaling >> Histones >> H4 >> Methylated
Applications
Show applications key
Our Abpromise guarantee covers the use of ab9053 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
ChIP/Chip: Use at an assay dependent concentration. PubMed: 19077033
IHC-P: Use at an assay dependent dilution. PubMed: 20007528
EM: Use at an assay dependent dilution. PubMed: 20543957
IHC (PFA fixed): Use at an assay dependent dilution. PubMed: 17083276
IHC-Fr: Use at an assay dependent dilution. PubMed: 17712411
ICC/IF: Use at an assay dependent dilution. PubMed: 19727073
Flow Cyt
Flow Cyt: 1/100.Flow Cyt: 1/100.
IF: 1/200 - 1/500.
WB: Use a concentration of 1 µg/ml. Detects a band of approximately 13 kDa (predicted molecular weight: 11 kDa).Can be blocked with Histone H4 peptide - tri methyl K20 (ab17567).
ChIP: Use 4-5µg for 106 cells.
Target
Relevance
Histone proteins H3, H4, H2A, and H2B function as building blocks to package eukaryotic DNA into repeating nucleosome units that are folded in higher order chromatin fibers. The nucleosome is composed of an octamer containing a H3/H4 tetramer and two H2A/H2B dimers, surrounded by approximately 146 base pairs of DNA. A diverse and elaborate array of post-translational modifications including acetylation, phosphorylation, methylation, ubiquitination, and ADP-ribosylation occurs on the N-terminal tail domains of histones. Methylation of position-specific lysine residues in histone N termini is a central modification for regulating epigenetic transitions in chromatin. Each methylatable lysine residue can exist in a mono, di, or tri methylated state. Arginine resdiues can also by mono or di methylated.
Cellular localization
Nuclear
Alternative names
- H4F2 antibody
- H4FN antibody
- HIST1H4 antibody
see all
Database links
- Entrez Gene: 8370 Human
- Entrez Gene: 121504 Human
- Entrez Gene: 320332 Mouse
- GenBank: NM_003548 Human
- Omim: 142750 Human
- SwissProt: P02304 Human
- SwissProt: P62805 Human
- SwissProt: P02304 Mouse
see all
Anti-Histone H4 (tri methyl K20) antibody - ChIP Grade images:
- Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053)
Left panel: a fission yeast whole cell extract and recombinant (r) fission yeast histone H4.
Right panel: fission yeast whole cell extracts from strains with only a single wt (h4) or mutant (h4K20R) gene were immunoblotted.
Review by Steven Sanders submitted 13 August 2004
ChIP - Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053)
Chromatin was prepared from U2OS cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 min. The ChIP was performed with 25 µg of chromatin, 6 µl of ab9053 (blue), and 20 µl 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). Primers and probes are located in the first kb of the transcribed region.
Western blot - Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053)
All lanes : Anti-Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053) at 1 µg/ml ab9053 is specific for Histone H4 (tri-methyl K20). This is illustrated in lane 5 where the activity of ab9053 is specifically blocked by the addition of the immunizing peptide (ab17567).
Lane 1 : Calf thymus histone lysate
Lane 2 : Calf thymus histone lysate with
Lane 3 : Calf thymus histone lysate with
Lane 4 : Calf thymus histone lysate with
Lane 5 : Calf thymus histone lysate with
Lane 6 : Calf thymus histone lysate with
Lane 7 : Calf thymus histone lysate with
Secondary
Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (HRP) (ab6721) at 1/5000 dilution
Predicted band size : 11 kDa
Observed band size : 13 kDa (why is the actual band size different from the predicted?)
Immunofluorescence - Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053)
HeLa cells immunofluorescently stained with anti-Tri Methyl Lysine 20 of histone H4 (ab9053). HeLa cells were cultured on glass coverslips, fixed with 4% paraformaldehyde, and immunofluorescently labeled with anti-Tri K20 (green) at 1/500 and DAPI (red).
This image is part of the nuclear antibody characterisation program at www.cellnucleus.com
The image was collected using a 1.3 N.A. 40 X PlanApo Objective (Zeiss) using a Zeiss Axiovert 200M inverted fluorescent microscope and a SensiCam HQ (Cooke Corp.) 12-bit CCD. The images were collected using Universal Imaging MetaMorph v. 5.1 for Windows. A Physik Instruments piezo z-drive was used to collect images at 200 nm intervals throughout the depth of the cell nucleus. The digital images were processed to remove fluorescent background and colour composites were made from single focal planes using MetaMorph
Michael Hendzel, Cross Cancer Inst, Edmonton, Canada
Immunofluorescence - Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053)
A mouse 10T1/2 embryonic fibroblast immunofluorescently stained with anti Tri Methyl Lysine 20 of histone H4. Murine 10T1/2 embryonic fibroblasts were cultured on glass coverslips, fixed with paraformaldehyde and then immunofluorescently labeled using an anti-Tri Methyl K20 of histone H4 antibody at 1/500 (green). The nuclear chromatin was simultaneously visualized using DAPI (red).
This image is part of the nuclear antibody characterisation program at www.cellnucleus.com
The image was collected using a 1.4 N.A. 100 X PlanApo Objective (Zeiss) using a Zeiss Axiovert 200M inverted fluorescent microscope and a Photometrics Cascade 16-bit CCD. The images were collected using Universal Imaging MetaMorph v. 5.1 for Windows. A Physik Instruments piezo z-drive was used to collect images at 200 nm intervals throughout the depth of the cell nucleus. The digital images were processed to remove fluorescent background and colour composites were made from single focal planes using MetaMorph.
Michael Henzel, Cross Cancer Inst, Edmonton, Canada
Immunofluorescence - Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053)
A field of HeLa cells stained with anti-Trimethy Lysine 20 of Histone H4.
This image is part of the nuclear antibody characterisation program at www.cellnucleus.com
Michael Henzel, Cross Cancer Inst, Edmonton, Canada
References for Anti-Histone H4 (tri methyl K20) antibody - ChIP Grade (ab9053)
This product has been referenced in:
- van Eggermond MC et al. Epigenetic regulation of CIITA expression in human T-cells. Biochem Pharmacol : (2011). ChIP; Human.Read more (PubMed: 21664896) »
- Poleshko A et al. Identification of a functional network of human epigenetic silencing factors. J Biol Chem 285:422-33 (2010). ChIP; Human.Read more (PubMed: 19880521) »
See all 43 publications for this product
Publishing research using ab9053? Please let us know so that we can cite the reference in this datasheet
Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"