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

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.

Monomethyl, dimethyl, and trimethyl histone H3 (Lys4). Additional unidentified bands above 50 kDa are detected in some samples

The immunizing sequence is conserved from Tetrahymena to human, so broad cross-reactivity is expected.

Epitope: Lys4 of Histone H3

KLH-conjugated, synthetic peptide containing …Tme2KQT… in which me2K corresponds to dimethyl-lysine at residue 4 of human histone H3.

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.

Research Category
Epigenetics & Nuclear Function

Research Sub Category
Histones

Use Anti-mono/di/trimethyl-Histone H3 (Lys4) Antibody, clone AW304 (rabbit monoclonal antibody) validated in ChIP, WB, PIA, Mplex to detect mono/di/trimethyl-Histone H3 (Lys4) also known as H3K4me, Histone H3 (methyl K4).

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.

~17 kDa

Replaces: 05-791

Cultured supernantant in 0.05% sodium azide

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.

Control
HeLa cell nuclear or acid extracts.

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