Western blot analysis of extracts from various cell lines using Symmetric Di-Methyl Histone H3 (Arg8) (E1W5H) Rabbit mAb.
The specificity of Symmetric Di-Methyl Histone H3 (Arg8) (E1W5H) Rabbit mAb was determined using peptide ELISA. The graph depicts the binding of the antibody to pre-coated, symmetric di-methyl histone H3 (Arg8) peptide in the presence of increasing concentrations of various competitor peptides. As shown, only the symmetric di-methyl histone H3 (Arg8) peptide competed away binding of the antibody.
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Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised June 2020
Protocol Id: 10
Symmetric Di-Methyl Histone H3 (Arg8) (E1W5H) Rabbit mAb recognizes endogenous levels of histone H3 protein only when symmetrically di-methylated at Arg8. This antibody may have a slight cross reactivity towards histone H3 protein when mono-methylated at Arg8.
Human, Monkey, Mouse, Rat
Bovine, S. cerevisiae, Zebrafish
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human histone H3 in which Arg8 is symmetrically di-methylated.
The nucleosome is the primary chromatin building block and consists of DNA wrapped around an octamer made of paired histone proteins H2A, H2B, H3, and H4. Chromatin remodeling plays a critical role in the regulation of various nuclear activities, including transcription. Histone proteins are targets of post-translational modification, including acetylation, phosphorylation, ubiquitination, and methylation. Modified histone residues are recognized and bound by chromatin modifiers and the transcription machinery to regulate gene expression (1-4). Protein arginine methyl transferases (PRMTs) methylate histone proteins at arginine residues to generate mono-methylated, symmetrically di-methylated, or asymmetrically di-methylated proteins. Asymmetrically di-methylated arginine residues are found on histone H3 (Arg2, 8, 17, 26 and 42), histone H4 (Arg3), and histone H2A (Arg3) proteins. Asymmetric methylation is carried out by type 1 PRMTs, which include PRMT1, PRMT2, PRMT4/CARM1, and PRMT6. These modifications are often associated with actively transcribed genes. Symmetric di-methylation of arginine residues are found on histone H3 (Arg2 and 8), Histones H4 (Arg3), and H2A (Arg3). Symmetrically di-methylated histone arginine residues are generated by type II transferases PRMT5 and PRMT7, and are often associated with transcription repression (5-9). Arginine residues can also be deiminated by a protein arginine deiminase (PADI) to form the non-coded amino acid citrulline. Conversion of arginine to citrulline prevents methylation of this residue and is thought to regulate histone arginine methylation levels (10-13).
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