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Products:Epigenetics and Nuclear Signaling >> Histones >> H3 >> Methylated
Overview
Product name
Anti-Histone H3 (mono methyl K4) antibody - ChIP Grade
See all Histone H3 products (41) ...
Description
Rabbit polyclonal to Histone H3 (mono methyl K4) - ChIP Grade
Specificity
Specific for mono-methylated Lysine 4 of histone H3. Does not recognise di- or tri-methyl Lysine 4 nor methylation at Lysine 9.
Tested applications
IP, Flow Cyt, Electron Microscopy, ICC/IF, ICC, ChIP, IF, WB, IHC-P, ChIP/Chipmore details
Cross reactivity
Reacts with
Human, Pig, Tetrahymena sp., Xenopus laevis
Predicted to work with
Cow, Indian Muntjac, Plants, all Mammals
Immunogen
Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3, mono methylated at K4.
(Peptide available as ab1340.)
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
pH: 7.40
Preservative: 0.02% Sodium azide
Constituent: PBS
Note: Batches of this product that have a concentration < 1mg/ml may have BSA added as a stabilising agent. If you would like information about the formulation of a specific lot, please contact our scientific support team who will be happy to help.
Concentration
Concentration information loading...
Purity
Immunogen affinity purified
Clonality
Polyclonal
Isotype
IgG
Research areas
Epigenetics and Nuclear Signaling >> ChIP'ing antibodies >> ChIP'ing antibodies
Epigenetics and Nuclear Signaling >> Histones >> H3 >> Methylated
Applications
Show applications key
Our Abpromise guarantee covers the use of ab8895 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
IP: Use at an assay dependent concentration.
Flow Cyt: 1/100
EM: Use at an assay dependent dilution. (PubMed: 20543957)
ICC/IF: Use at an assay dependent dilution.
ICC: Use at an assay dependent concentration.
ChIP: Use 2 µg for 25 µg of chromatin.
IF
IF: 1/500IF: 1/500
WB: 1/500Detects a band of approximately 17 kDa (predicted molecular weight: 15 kDa).Can be blocked with Histone H3 peptide - mono methyl K4 (ab1340).
IHC-P: Use at an assay dependent dilution.
ChIP/Chip: Use at an assay dependent dilution.
Target
Function
Core component of nucleosome. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling.
Sequence similarities
Belongs to the histone H3 family.
Developmental stage
Expressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
Post-translational
modifications
Acetylation is generally linked to gene activation. Acetylation on Lys-10 (H3K9ac) impairs methylation at Arg-9 (H3R8me2s). Acetylation on Lys-19 (H3K18ac) and Lys-24 (H3K24ac) favors methylation at Arg-18 (H3R17me).
Citrullination at Arg-9 (H3R8ci) and/or Arg-18 (H3R17ci) by PADI4 impairs methylation and represses transcription.
Asymmetric dimethylation at Arg-18 (H3R17me2a) by CARM1 is linked to gene activation. Symmetric dimethylation at Arg-9 (H3R8me2s) by PRMT5 is linked to gene repression. Asymmetric dimethylation at Arg-3 (H3R2me2a) by PRMT6 is linked to gene repression and is mutually exclusive with H3 Lys-5 methylation (H3K4me2 and H3K4me3). H3R2me2a is present at the 3' of genes regardless of their transcription state and is enriched on inactive promoters, while it is absent on active promoters.
Methylation at Lys-5 (H3K4me), Lys-37 (H3K36me) and Lys-80 (H3K79me) are linked to gene activation. Methylation at Lys-5 (H3K4me) facilitates subsequent acetylation of H3 and H4. Methylation at Lys-80 (H3K79me) is associated with DNA double-strand break (DSB) responses and is a specific target for TP53BP1. Methylation at Lys-10 (H3K9me) and Lys-28 (H3K27me) are linked to gene repression. Methylation at Lys-10 (H3K9me) is a specific target for HP1 proteins (CBX1, CBX3 and CBX5) and prevents subsequent phosphorylation at Ser-11 (H3S10ph) and acetylation of H3 and H4. Methylation at Lys-5 (H3K4me) and Lys-80 (H3K79me) require preliminary monoubiquitination of H2B at 'Lys-120'. Methylation at Lys-10 (H3K9me) and Lys-28 (H3K27me) are enriched in inactive X chromosome chromatin.
Phosphorylated at Thr-4 (H3T3ph) by GSG2/haspin during prophase and dephosphorylated during anaphase. Phosphorylation at Ser-11 (H3S10ph) by AURKB is crucial for chromosome condensation and cell-cycle progression during mitosis and meiosis. In addition phosphorylation at Ser-11 (H3S10ph) by RPS6KA4 and RPS6KA5 is important during interphase because it enables the transcription of genes following external stimulation, like mitogens, stress, growth factors or UV irradiation and result in the activation of genes, such as c-fos and c-jun. Phosphorylation at Ser-11 (H3S10ph), which is linked to gene activation, prevents methylation at Lys-10 (H3K9me) but facilitates acetylation of H3 and H4. Phosphorylation at Ser-11 (H3S10ph) by AURKB mediates the dissociation of HP1 proteins (CBX1, CBX3 and CBX5) from heterochromatin. Phosphorylation at Ser-11 (H3S10ph) is also an essential regulatory mechanism for neoplastic cell transformation. Phosphorylated at Ser-29 (H3S28ph) by MLTK isoform 1, RPS6KA5 or AURKB during mitosis or upon ultraviolet B irradiation. Phosphorylation at Thr-7 (H3T6ph) by PRKCBB is a specific tag for epigenetic transcriptional activation that prevents demethylation of Lys-5 (H3K4me) by LSD1/KDM1A. At centromeres, specifically phosphorylated at Thr-12 (H3T11ph) from prophase to early anaphase, by DAPK3 and PKN1. Phosphorylation at Thr-12 (H3T11ph) by PKN1 is a specific tag for epigenetic transcriptional activation that promotes demethylation of Lys-10 (H3K9me) by KDM4C/JMJD2C. Phosphorylation at Tyr-42 (H3Y41ph) by JAK2 promotes exclusion of CBX5 (HP1 alpha) from chromatin.
Monoubiquitinated by RAG1 in lymphoid cells, monoubiquitination is required for V(D)J recombination (By similarity). Ubiquitinated by the CUL4-DDB-RBX1 complex in response to ultraviolet irradiation. This may weaken the interaction between histones and DNA and facilitate DNA accessibility to repair proteins.
Cellular localization
Nucleus. Chromosome.
Target information above from: UniProt accessionP68431
The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010).
Alternative names
- Histone H3.1 antibody
- FLJ92264 antibody
- H3 histone antibody
see all
Database links
- Entrez Gene: 326601 Cow
- Entrez Gene: 8350 Human
- Entrez Gene: 8351 Human
- Entrez Gene: 8352 Human
- Entrez Gene: 8353 Human
- Entrez Gene: 8354 Human
- Entrez Gene: 8355 Human
- Entrez Gene: 8356 Human
see all
Anti-Histone H3 (mono methyl K4) antibody - ChIP Grade images:
Western blot
All lanes : Anti-Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895) at 1/500 dilution
Lane 1 : Calf thymus histone lysate
Lane 2 : Calf thymus histone lysate with Histone H3 peptide - mono methyl K4 (ab1340) at 1 µg/ml
Lane 3 : Calf thymus histone lysate with Histone H3 peptide - di methyl K4 (ab7768) at 1 µg/ml
Lane 4 : Calf thymus histone lysate with Histone H3 peptide - tri methyl K4 (ab1342) at 1 µg/ml
Lane 5 : Calf thymus histone lysate with Histone H3 peptide - mono methyl K9 (ab1771) at 1 µg/ml
Lane 6 : Calf thymus histone lysate with Histone H3 peptide - mono methyl K27 (ab1780) at 1 µg/ml
Lane 7 : Calf thymus histone lysate with Histone H3 peptide - unmodified (ab2903) at 1 µg/ml
Secondary
Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (HRP) (ab6721) at 1/5000 dilution
Performed under reducing conditions.
Predicted band size : 15 kDa
Observed band size : 18 kDa (why is the actual band size different from the predicted?)
Exposure time : 2 minutes
ab8895 is specific for mono-methylated Lysine 4 of histone H3 and does not recognize di- or tri-methyl Lysine 4 nor methylation at Lysine 9. This is shown in lane 2 where the activity of the antibody is specifically blocked by the addition of the immunizing peptide (ab1340).
ChIP - Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895)
Chromatin was prepared from U2OS cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10min. The ChIP was performed with 25µg of chromatin, 2µg of ab8895 (blue), and 20µl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified on the GAPDH and ALDOA (active) and MYO-D (inactive) promoters and over the y-Actin gene (active). Schematic diagram of the y-Actin gene is shown on the top of the figure. Black boxes represent exons and thin lines represent introns. PCR products are depicted as bars under the gene.
Electron Microscopy - Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895)
A: Enlarged region showing the distribution of the H3K4me1 labelling around and within the EC cavities of a murine rod photoreceptor.B: Schematic representation of the labelling for the H3K4me1 mark. The bar represents 260 nm in A and 1 µm in B.
Image from Kizilyaprak C et al, PLoS One. 2010 Jun 9;5(6):e11039, Fig 6.
Dot Blot - Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895)
ab8895 used in Dot Blot at a 1/1000 dilution, 18 hours at 4°C.The dot blot was done using 0.1ug of peptide and screened multiple histone tail modifications to ensure no cross reactivity.
Image courtesy of an anonymous Abreview.
Immunocytochemistry/ Immunofluorescence - Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895)
Indian muntjac fibroblast cells - interphase (top left) and prophase (top right and below), stained with:
Mono Methyl K4 antibody, ab8895, (green)
DAPI: red, top left and right; blue, below
Phospho Ser 10 antibody: red (below) and blue (top right)
The perinuclear and perinucleolar heterochromatin domains do not contain Mono Methyl K4. The Mono Methyl K4, rather, is distributed as small nuclear foci primarily found between DAPI-intense regions of the nucleus.
Kirk McManus in the lab of Michael Hendzel, Univeristy of Alberta
Flow Cytometry - Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895)
ab8895 staining mouse embryonic stem cells by flow cytometry. The ES cell colonies were trypsinized and the cells permeabilized before being stained with ab8895 (0.25ug/1.5 x 105 cells). A PE conjugated goat anti-rabbit antibody was used as the secondary.
This image is courtesy of an Abreview submitted by Prof Albrecht Müller
Protocols
References for Anti-Histone H3 (mono methyl K4) antibody - ChIP Grade (ab8895)
This product has been referenced in:
- Kwon Tet al. Mechanism of histone lysine methyl transfer revealed by the structure of SET7/9-AdoMet. EMBO J 22:292-303 (2003).Read more (PubMed: 12514135) »
- Medyouf Het al. High-level IGF1R expression is required for leukemia-initiating cell activity in T-ALL and is supported by Notch signaling. J Exp Med 208:1809-22 (2011). ChIP.Read more (PubMed: 21807868) »
See all 71 publications for this product
Publishing research using ab8895? 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"