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ChIP - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
![ChIP - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)](/ps/datasheet/images/ab6002_9.jpg)
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Western blot - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
![Western blot - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)](/ps/datasheet/images/ab6002 copy.jpg)
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Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
![Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)](/ps/datasheet/images/ab6002_4.jpg)
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Product Name
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Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade
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See all Histone H3 (methylated) antibodies (36)...
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Product type
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Primary antibodies
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Description
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Mouse monoclonal [mAbcam 6002] to Histone H3 (tri methyl K27) - ChIP Grade
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Immunogen
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Synthetic peptide derived from residues 1 - 100 of Human Histone H3, tri methylated at K27.
(Note: the amino acid sequence is proprietar y).
Hybridoma were prepared and the resulting clones were positively screened by ELISA against the immunising peptide. Clones were negatively screened against both the corresponding unmodified peptide and also against a peptide corresponding to tri methylated K9 of Histone H3.
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Reacts with
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Hu, Ms, Cow, Dm, Plnts, Zfsh
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Predicted to react
(species key)
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Rat, Chk, ChHm, Rb, Xl
(Due to sequence homology)
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Specificity
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This antibody is specific for histone H3 tri-methylated at K27.
The sequence is found in all mammals and a wide range of species, including D. melanogaster, Arabidopsis, Chicken and Xenopus.
The antibody will react with any of the above species where the modification is present. Reactivity was confirmed using Cow samples.
Reactivity is not certain in S. pombe and S. cerevisiae as the equivalent protein sequence differs slightly from the species listed above. A customer reported that the activity against histone H3 in Zea mays (maize) is not good.
The antibody is blocked in western blotting by tri methyl K27 peptide and slightly by di methyl K27 peptide. It is not blocked by mono methyl K4, di methyl K4, tri methyl K4, mono methyl K9, di methyl K9, tri methyl K9, mono methyl K27 or unmodified K27 peptides.
(See blot below).
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Tested applications
(see key)
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ChIP, ChIP/Chip, Flow Cyt, ICC/IF, IHC-P, WB
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Abreviews
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Application notes
(see key)
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Recommended dilutions
ChIP: Use 5-10µg for 25µg of chromatin
ChIP on chip: Use at an assay dependent dilution.
ICC/IF: Use at an assay dependent concentration.
WB: Use at a concentration of 3 - 5 µg/ml. Detects a band of approximately 17 kDa (predicted molecular weight: 15 kDa). Can be blocked with Histone H3 peptide - tri methyl K27 (ab1782).
IHC-P: Use at an assay dependent concentration.
Flow Cyt: Use at an assay dependent concentration.
Not yet tested in other applications.
Optimal dilutions/concentrations should be determined by the end user.
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Cellular localization
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Nuclear
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Research areas
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Chromatin and Nuclear Signaling >> ChIP'ing antibodies >> ChIP'ing antibodies
Chromatin and Nuclear Signaling >> Histones >> H3 >> Methylated
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Relevance
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Histones are basic nuclear proteins that, together with DNA, make up the nucleosome structure in eukaryotes. Nucleosomes consist of approximately 146 bp of DNA wrapped around a histone octamer, which is composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin structure is further compacted through the interaction of a linker histone H1.
Methylation of specific residues in the histone tails is a central modification for regulating epigenetic transitions in chromatin. Modified lysine residues can exist in a mono-, di-, or tri-methylated state, whilst arginine residues can be mono- or di-methylated.
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Alternative names Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002) |
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Database links
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The links below go to external sites and will open in a new browser window |
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Raised in
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Mouse
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Clonality
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Monoclonal
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Clone number
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mAbcam 6002
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Isotype
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IgG3
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Light chain type
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kappa
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Purity
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Protein G purified
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Storage buffer
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Preservative: 0.02% Sodium Azide
Constituents: PBS, pH 7.4 Material safety datasheet (MSDS) for this product:
Sodium Azide MSDS
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Form
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Liquid
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Concentration
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Batch dependent within range: 0.90 - 1.00 mg/ml
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Storage instructions
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Store at +4°C short term (1-2 weeks). Aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
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At Abcam, we have one centralized database to hold all of our product information, so that everything we know about this Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade is on this datasheet. But please do contact us if you would like any reassurance! |
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See below for Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade images, references, products related to ab6002 and other tools.
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Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade images:
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 ChIP - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
Chromatin was prepared from Hela cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min. The ChIP was performed with 25µg of chromatin, 5µg of ab6002 (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.
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Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
Immunofluorescent imaging of human cells (U2OS) with ab6002 reveals broadly dispersed interphase nuclear staining corresponding to trimethylation of K27, with multiple foci of brighter staining, exactly agreeing with published studies of K27-trimethyl IF [See figure 3 of Peters et al Mol Cell 12(6):1577-1589 (2003)] . The lack of nucleolar or cytoplasmic staining background confirms the high specificity of the antibody in this application.
IF was performed with a standard paraformaldehyde technique (fixed in PBS buffered PFH 4% for 5 minutes, permeabilised with 0.5% triton-PBS for 5 minutes, blocked with 5% milk / 0.2% tween for one hour. Primary antibody used at 1/200 in 5% milk / 0.2% TWEEN for one hour, secondary antibody Alexa 488 for 30 minutes. All blocking and incubation steps carried out at 37 degrees C.
Luke Hughes-Davies and Rhiannon Jade, Gurdon Institute, Cambridge, UK
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Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
Staining of facultative heterochromatin of inactive X chromosomes
![Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)](/ps/datasheet/images/ab6002_51.jpg)
Figure showing the nuclear distribution of H3 (tri-methyl K27) antibody, ab6002 in a) a 46 chromosome, XX cell line, and b) a 49 chromosome, XXXXX cell line.
The location of facultative heterochromatin at the inactive X chromosome is indicated by white arrow heads.
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Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
![Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)](/ps/datasheet/images/ab6002_6.jpg)
Interphase 10T1/2 mouse fibroblasts were paraformaldehyde fixed (4%), immunofluorescently labeled with anti-trimethyl K27 antibody (ab6002) and counterstained with DAPI. The merge image presents the DAPI and ab6002 channels as red and green, respectively. The scale bar represents 3µm. Kirk Mcmanus
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- Histone H3 (tri methyl K27) antibody [mAbcam 6002] - IF Grade (ab6002)
![- Histone H3 (tri methyl K27) antibody [mAbcam 6002] - IF Grade (ab6002)](/ps/datasheet/images/ab6002_9.gif)
ELISA using ab6002 at antibody concentrations between 1/500 and 1/8000.
The red line indicates binding to the tri methyl K27 peptide (ab1782). Binding to the following peptides was not seen:
Unmodified K27 (ab2623),
mono methyl K27 (ab1780),
di methyl K27 (ab1781),
mono methyl K9 (ab1771),
di methyl K9 (ab1772),
tri methyl K9 (ab1773),
mono methyl K4 (ab1340),
di methyl K4 (ab7768),
tri methyl K4 (ab1342).
This indicates the specificity of ab6002 for tri methyl K27 of Histone H3.
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Flow Cytometry - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
![Flow Cytometry - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)](/ps/datasheet/images/ab6002_10.jpg)
ab6002 staining mouse embyronic stem cells by flow cytometry. The ES cell colonies were trypsinized and permeabilized prior to blocking and staining with the antibody (1ug/1.5 x 10 5 cells. A Cy3 conjugated anti-mouse antibody was used as the secondary. This image is courtesy of an Abreview submitted by Prof Albrecht Müller
See Abreview
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Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
![Immunocytochemistry/ Immunofluorescence - Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)](/ps/datasheet/images/6/ab6002/Histone-H3-Primary-antibodies-ab6002-6.jpg)
ICC/IF image of ab6002 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab6002, 5µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-mouse IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM. This antibody also gave a positive result in 4% PFA fixed (10 min) Hek293, HepG2 and MCF7 cells at 5µg/ml, and in 100% methanol fixed (5 min) HeLa, Hek293, HepG2 and MCF7 cells at 5µg/ml.
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References for Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade (ab6002)
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This product has been used in: (two most recent references)
Verdeguer F et al. A mitotic transcriptional switch in polycystic kidney disease. Nat Med 16:106-10 (2010). ChIP; Mouse. PubMed: 19966811
Obier N & Mller AM Chromatin flow cytometry identifies changes in epigenetic cell states. Cells Tissues Organs 191:167-74 (2010). Flow Cyt; Mouse. PubMed: 19776550
See all 38 publication references for this product.
If you publish research using ab6002 please let us know so that we can cite the reference on this datasheet.
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Search PubMed (MEDLINE) for references to Histone H3
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Histone H3 (tri methyl K27) antibody [mAbcam 6002] - ChIP Grade - more information
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All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"
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