Anti-Histone H3 (methylated) antibody [mAbcam 1220] - ChIP Grade (ab1220)

Overview

• Product nameAnti-Histone H3 (methylated) antibody [mAbcam 1220] - ChIP GradeSee all Histone H3 (methylated) primary antibodies ...
• Description
  Mouse monoclonal [mAbcam 1220] to Histone H3 (methylated) - ChIP Grade
• SpecificityBy peptide ELISA ab1220 recognises di methyl K9, but not unmodified K9, mono methyl K9, tri methyl K9, di methyl K27, tri methyl K27, mono methyl K4, di methyl K4 or tri methyl K4. By Western blot ab1220 is blocked by di methyl K9, but not by unmodified K9, mono methyl K9, tri methyl K9, di methyl K27, tri methyl K27, mono methyl K4, di methyl K4 or tri methyl K4. This indicates the specificity of ab1220 for di methyl K9 of Histone H3.
• Tested applicationsICC/IF, IP, WB, Flow Cyt, IHC-Fr, ELISA, IHC-P, ChIP more details
• Species reactivity
  Reacts with: Mouse, Rat, Chicken, Cow, Human, Xenopus laevis, Arabidopsis thaliana, Caenorhabditis elegans, Fruit fly (Drosophila melanogaster), Schizosaccharomyces pombe, Corn, Marmoset (common), Rice, Other
  Predicted to work with: Sheep, Saccharomyces cerevisiae
• Immunogen

  Synthetic peptide conjugated to KLH derived from within residues 1 - 100 of Human Histone H3, di methylated at K9.

  Read Abcam's proprietary immunogen policy

  . Hybridomas were prepared and the resulting clones were positively screened by ELISA against the immunising peptide conjugated to BSA. Clones were negatively screened against both the non-modified equivalent peptide and against a peptide corresponding to dimethylated K27.

Properties

• FormLiquid
• Storage instructionsStore at +4°C short term (1-2 weeks). Aliquot and store at -20°C or -80°C. Avoid repeated freeze / thaw cycles.
• Storage bufferpH: 7.50
  Preservative: 0.02% Sodium azide
  Constituent: PBS
  
  Some batches of this product contain 0.4M Arginine 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 information loading...
• PurityIgG fraction
• Clonality Monoclonal
• Clone numbermAbcam 1220
• IsotypeIgG2a
• Light chain typekappa
• Research Areas
  • Epigenetics and Nuclear Signaling
  • Histones
  • H3
  • Methylated

  • Epigenetics and Nuclear Signaling
  • ChIP'ing antibodies
  • ChIP'ing antibodies

Applications

Target

• FunctionCore 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 similaritiesBelongs to the histone H3 family.
• Developmental stageExpressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
• Post-translational
  modificationsAcetylation 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 localizationNucleus. Chromosome.

Target information above from: UniProt accession P68431 The UniProt Consortium
The Universal Protein Resource (UniProt) in 2010
Nucleic Acids Res. 38:D142-D148 (2010) .

Information by UniProt
• Database links
  • Entrez Gene: 176359 Caenorhabditis elegans
  • Entrez Gene: 427887 Chicken
  • 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
  • Entrez Gene: 8357 Human
  • Entrez Gene: 8358 Human
  • Entrez Gene: 8968 Human
  • Entrez Gene: 319152 Mouse
  • Entrez Gene: 319153 Mouse
  • Entrez Gene: 360198 Mouse
  • Entrez Gene: 97908 Mouse
  • Entrez Gene: 100364501 Rat
  • Entrez Gene: 100365669 Rat
  • Entrez Gene: 291159 Rat
  • Entrez Gene: 679994 Rat
  • Entrez Gene: 680511 Rat
  • Entrez Gene: 682330 Rat
  • Omim: 601128 Human
  • SwissProt: P59226 Arabidopsis thaliana
  • SwissProt: Q10453 Caenorhabditis elegans
  • SwissProt: P08898 Caenorhabditis elegans
  • SwissProt: P84229 Chicken
  • SwissProt: P68432 Cow
  • SwissProt: P02299 Fruit fly (Drosophila melanogaster)
  • SwissProt: Q93081 Human
  • SwissProt: Q16695 Human
  • SwissProt: P68431 Human
  • SwissProt: P68433 Mouse
  • SwissProt: Q6LED0 Rat
  • Unigene: 132854 Human
  • Unigene: 247813 Human
  • Unigene: 247814 Human
  • Unigene: 248176 Human
  • Unigene: 443021 Human
  • Unigene: 484990 Human
  • Unigene: 532144 Human
  • Unigene: 533292 Human
  • Unigene: 546315 Human
  • Unigene: 586261 Human
  • Unigene: 591778 Human
  • Unigene: 221301 Mouse
  • Unigene: 261657 Mouse
  • Unigene: 377874 Mouse
  • Unigene: 390558 Mouse
  • Unigene: 397328 Mouse
  • Unigene: 138090 Rat

  see all

• Alternative names
  FLJ92264 antibodyH3 histone antibodyH3 histone antibody

  H3 histone family, member A antibodyH3/a antibodyH3/b antibodyH3/c antibodyH3/d antibodyh3/f antibodyH3/h antibodyH3/i antibodyH3/j antibodyH3/k antibodyH3/l antibodyH31_HUMAN antibodyH3F1K antibodyH3F3 antibodyH3FA antibodyH3FB antibodyH3FC antibodyH3FD antibodyH3FF antibodyH3FH antibodyH3FI antibodyH3FJ antibodyH3FK antibodyH3FL antibodyHIST1H3A antibodyHIST1H3B antibodyHIST1H3C antibodyHIST1H3D antibodyHIST1H3E antibodyHIST1H3F antibodyHIST1H3G antibodyHIST1H3H antibodyHIST1H3I antibodyHIST1H3J antibodyHIST3H3 antibodyHistone 1, H3a antibodyHistone cluster 1, H3a antibodyHistone cluster 1, H3b antibodyHistone cluster 1, H3c antibodyHistone cluster 1, H3d antibodyHistone cluster 1, H3e antibodyHistone cluster 1, H3f antibodyHistone cluster 1, H3g antibodyHistone cluster 1, H3i antibodyHistone cluster 1, H3j antibodyHistone H 3 antibodyHistone H3.1 antibodyHistone H3.1 antibodyHistone H3/a antibodyHistone H3/b antibodyHistone H3/c antibodyHistone H3/d antibodyHistone H3/f antibodyHistone H3/h antibodyHistone H3/i antibodyHistone H3/j antibodyHistone H3/k antibodyHistone H3/l antibody

  see all

Anti-Histone H3 (methylated) antibody [mAbcam 1220] - ChIP Grade images

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Histone H3 (di methyl K9) antibody [mAbcam 1220] - ChIP Grade (ab1220)

  ab1220 staining human kidney sections by IHC-P using EXPOSE IHC detection kit (ab80436). Formalin fixed paraffin embedded tissue sections were pre-treated using heat mediated antigen retrieval (using a pressure cooker) with sodium citrate buffer (pH6) for 30 mins. The section was incubated with ab1220, 5µg/ml, for 1 hour at room temperature. DAB was used as the chromogen and the section was counterstained with haematoxylin and mounted with DPX.

• 

  Western blot
  All lanes : Anti-Histone H3 (methylated) antibody [mAbcam 1220] - ChIP Grade (ab1220)
  
  Lane 1 : Calf thymus histone lysate
  Lane 2 : Calf thymus histone lysate with Histone H3 peptide - unmodified at 1 µg/ml
  Lane 3 : Calf thymus histone lysate with Histone H3 peptide - mono methyl K9 (ab1771) at 1 µg/ml
  Lane 4 : Calf thymus histone lysate with Histone H3 peptide - di methyl K9 (ab1772) at 1 µg/ml
  Lane 5 : Calf thymus histone lysate with Histone H3 peptide - tri methyl K9 (ab1773) at 1 µg/ml
  Lane 6 : Calf thymus histone lysate with Histone H3 peptide - di methyl K4 (ab7768) at 1 µg/ml
  Lane 7 : Calf thymus histone lysate with Histone H3 peptide - di methyl K27 (ab1781) at 1 µg/ml
  
  Secondary
  Rabbit polyclonal Secondary Antibody to Mouse IgG - H&L (HRP) (ab6728) at 1/5000 dilution
  developed using the ECL technique
  
  Performed under reducing conditions.
  
  Predicted band size : 17 kDa
  
  
  Exposure time : 1 minute
• 

  ChIP - Histone H3 (di methyl K9) antibody [mAbcam 1220] - ChIP Grade (ab1220)

  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  ab1220 (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.
   

• 

  Immunocytochemistry/ Immunofluorescence - Histone H3 (di methyl K9) antibody [mAbcam 1220] - ChIP Grade (ab1220)This image was submitted as part of a review by Kirk McManus

  Interphase HeLa cells were fixed in paraformaldehyde and labelled with ab1220 (1/500). The staining is shown in green. The cells were counterstained with DAPI (red).

• 

  - Histone H3 (di methyl K9) antibody [mAbcam 1220] (ab1220)

  ELISA using ab1220 at varying antibody concentrations.

  The purple line indicates binding to the di methyl K9 peptide ab1772. Binding to the following peptides was not seen:
  unmodified K9 (ab2903),
  mono methyl K9 (ab1771),
  tri methyl K9 (ab1773),
  di methyl K27 (ab1781),
  tri methyl K27 peptide (ab1782),
  mono methyl K4 (ab1340),
  di methyl K4 (ab7768),
  tri methyl K4 (ab1342).

  This indicates the specificity of ab1220 for di methyl K9 of Histone H3.
  

• 

  Flow Cytometry - Histone H3 (di methyl K9) antibody [mAbcam 1220] - ChIP Grade (ab1220)This image is courtesy of an Abreview submitted by Prof Albrecht Müller

  ab1220 staining mouse embryonic stem cells by flow cytometry (gated on all living cells). The cells were trypsinized and stained with the antibody at 1ug/1.5 x 10⁵ cells in a permeabilization buffer. A Cy3 ® conjugated goat anti-mouse antibody was used as the secondary.

  See Abreview

• 

  Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Histone H3 (di methyl K9) antibody [mAbcam 1220] - ChIP Grade (ab1220)This image is courtesy of an anonymous Abreview

  ab1220 staining rat liver tissue sections by IHC-P.  Sections were formaldehyde fixed and subjected to heat mediated antigen retrieval in citrate buffer pH 6.0 prior to blocking with 5% serum for 30 minutes at 20°C.  The primary antibody was diluted 1/400 and incubated with the sample for 45 minutes at 20°C.  A HRP-conjugated goat anti-mouse antibody was used as the secondary.

  See Abreview