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  • : 04-791
  • : 100 μL
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Peptide Inhibition Analysis
Representative blot from a previous lot. Acid extracts from HeLa cells were resolved by electrophoresis, transferred ...

Beadlyte® Histone-Peptide Specificity Assay
1:1,000 to 1:2,000 dilutions of a previous lot were incubated with a cocktail of microspheres conjuga...

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Anti-mono/di/trimethyl-Histone H3 (Lys4) Antibody, clone AW304

Recommended Replacement for: 05-791
Species Reactivity Key Applications Host Format Antibody Type
H, Vrt WB, PIA, Mplex Rabbit null Monoclonal Antibody
Anti-mono/di/trimethyl-Histone H3 (Lys4) Antibody, clone AW304 | 04-791
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Trade Name:
Upstate (Millipore)
Monomethyl, dimethyl, and trimethyl histone H3 (Lys4). Additional unidentified bands above 50 kDa are detected in some samples
Molecular Weight:
~17 kDa
Lys4 of Histone H3
KLH-conjugated, synthetic peptide containing …Tme2KQT… in which me2K corresponds to dimethyl-lysine at residue 4 of human histone H3.
Background Information:
Histones are highly conserved proteins that serve as the structural scaffold for the organization of nuclear DNA into chromatin. The four core histones, H2A, H2B, H3, and H4, assemble into an octamer (2 molecules of each). Subsequently, 146 base pairs of DNA are wrapped around the octamer, forming a nucleosome, the basic subunit of chromatin. Histones are modified post-translationally by the actions of enzymes in both the nucleus and cytoplasm. These modifications regulate DNA transcription, repair, recombination, and replication. The most commonly studied modifications are acetylation, phosphorylation, methylation, and ubiquitination. These modifications can alter local chromatin architecture, or recruit trans-acting factors that recognize specific histone modifications (the "histone code" hypothesis). The modifications occur predominantly on the N-terminal and C-terminal tails that extend beyond the nucleosome core particle. Histone methylation is primarily associated with heterochromatin and gene silencing.
Species Reactivity:
  • Human
  • Vertebrates
Species Reactivity Note:
Human. The immunizing sequence is conserved from Tetrahymena to human, so broad cross-reactivity is expected.
Application Notes:
Chromatin Immunoprecipitation:
The antibody was reported by an independent laboratory as suitable for ChIP.

Immunoblot Analysis:
A previous lot detected methylated Histone H3 in acid-extracted proteins from HeLa cells. The antibody did not detect unmethylated recombinant histone H3 (Catalog # 14-494).

Peptide Inhibition Analysis:
2 μM of a histone H3 peptide containing monomethyl-lysine 4, dimethyl lysine 4, and trimethyl-lysine 4 abolished detection of histone H3 by this lot in immunoblot analysis of HeLa acid extracts. Some signal reduction was observed with histone H3 peptides containing dimethyl-lysine 9 or 27.

Beadlyte® Histone-Peptide Specificity Assay: 1:1,000-1:2,000 dilutions of this lot were incubated with histone H3 peptides containing various modifications conjugated to Luminex® microspheres. Specificity for mono, di, and trimethyl-lysine 4 was demonstrated.
HeLa cell nuclear or acid extracts.
Quality Assurance:
Routinely evaluated by immunoblot on nuclear extract from HeLa cells.

Immunoblot Analysis: A 1:500 (lane 1) and 1:1000 (lane 2) dilution of this lot detected methylated Histone H3 in HeLa nuclear extracts.
Cultured supernantant in 0.05% sodium azide
Storage Conditions:
Stable for 1 year at 2-8°C from date of receipt.
For maximum recovery of product, centrifuge the vial prior to removing the cap. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance.
UniProt Number:
Entrez Gene Number:
Gene Symbol:
  • HIST3H3
  • H3FT
  • MGC126886
  • H3t
  • MGC126888
  • H3T
  • H3/g
  • H3.4
  • H3/t
Alternate Names:
  • H3K4me
  • Histone H3 (methyl K4)
Usage Statement:
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Key Applications:
  • Western Blotting
  • Peptide Inhibition Assay
  • Multiplexing
Entrez Gene Summary:
Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Nucleosomes consist of approximately 146 bp of DNA wrapped around a histone octamer composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin fiber is further compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures. This gene is intronless and encodes a member of the histone H3 family. Transcripts from this gene lack polyA tails; instead, they contain a palindromic termination element. This gene is located separately from the other H3 genes that are in the histone gene cluster on chromosome 6p22-p21.3.
UniProt Summary:
FUNCTION: SwissProt: Q16695 # 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.
SIZE: 136 amino acids; 15508 Da
SUBUNIT: The nucleosome is a histone octamer containing two molecules each of H2A, H2B, H3 and H4 assembled in one H3-H4 heterotetramer and two H2A-H2B heterodimers. The octamer wraps approximately 147 bp of DNA.
TISSUE SPECIFICITY: Expressed in testicular cells.
DEVELOPMENTAL STAGE: Expressed during S phase, then expression strongly decreases as cell division slows down during the process of differentiation.
PTM: Acetylation is generally linked to gene activation. Acetylation on Lys-10 impairs methylation at Arg-9. Acetylation on Lys-19 and Lys-24 favors methylation at Arg-18 (By similarity). & Citrullination at Arg-9 and/or Arg-18 by PADI4 impairs methylation and represses transcription (By similarity). & Asymmetric dimethylation at Arg-18 by CARM1 is linked to gene activation. Symmetric dimethylation at Arg-9 by PRMT5 is linked to gene repression (By similarity). & Methylation at Lys-5, Lys-37 and Lys-80 are linked to gene activation. Methylation at Lys-5 facilitates subsequent acetylation of H3 and H4. Methylation at Lys-80 is associated with DNA double-strand break (DSB) responses and is a specific target for TP53BP1. Methylation at Lys-10 and Lys-28 are linked to gene repression. Methylation at Lys-10 is a specific target for HP1 proteins (CBX1, CBX3 and CBX5) and prevents subsequent phosphorylation at Ser-11 and acetylation of H3 and H4. Methylation at Lys-5 and Lys-80 require preliminary monoubiquitination of H2B at 'Lys-120'. Methylation at Lys-10 and Lys-28 are enriched in inactive X chromosome chromatin (By similarity). & Phosphorylated at Thr-4 by GSG2/haspin during prophase and dephosphorylated during anaphase. At centromeres, specifically phosphorylated at Thr-12 from prophase to early anaphase. Phosphorylated at Ser-11 during the whole mitosis. Phosphorylation at Ser-11, which is linked to gene activation, prevents methylation at Lys-10 but facilitates acetylation of H3 and H4. Phosphorylated at Ser-29 by MLTK isoform 1, RPS6KA5 or AURKB during mitosis or upon ultraviolet B irradiation (By similarity). & Phosphorylation at 'Ser-11' is crucial for chromosome condensation and cell-cycle progression during mitosis and meiosis. In addition phosphorylation at 'Ser-11' is important during interphase because it enables the transcription of genes following external stimulation, like stress or growth factors. Phosphorylation at 'Ser-11' is also an essential regulatory mechanism for neoplastic cell transformation. Phosphorylation at 'Ser-11' by AURKB/Aurora-B mediates the dissociation of HP1 proteins (CBX1, CBX3 and CBX5) from heterochromatin. & Ubiquitinated (By similarity).
SIMILARITY: SwissProt: Q16695 ## Belongs to the histone H3 family.
Brand Family:
Product Name:
Anti-mono/di/trimethyl-Histone H3 (Lys4), clone AW304
Antibody Type:
Monoclonal Antibody
100 μL

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