Anti-monomethyl Histone H4 (Lys20) Antibody, Trial Size

Histones are highly conserved proteins that serve as the structural scaffold for the organization of nuclear DNA into chromatin. Histones 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). Methylation of histone H4 at lysine 20 has been implicated in transcriptional activation, gene silencing, heterochromatin formation, mitosis, and DNA repair. Histone H4 is progressively methylated at Lys20 during the G(2), M, and G(1) phases of the cell cycle, and methylation of H4Lys20 is a marker for heterochromatin. In mammals, the female X chromosome is coated by the Xist RNA, which is critical for silencing, and is marked mainly by methylation of H3Lys9, H3Lys27 and H4Lys20.

~11 kDa

Epitope: Monomethyl Lys20

KLH-conjugated, linear synthetic peptide corresponding to amino acids 15-24
(AKRHR[me1K]VLRD-C) of human Histone H4.

Anti-monomethyl Histone H4 (Lys20) Antibody, Trial Size is a rabbit polyclonal antibody for detection of monomethyl Histone H4 (Lys20) also known as H4K20me1, Histone H4 (mono methyl K20), H4 histone family member A & has been validated in WB.

Chromatin Immunoprecipitation:
Sonicated chromatin prepared from HeLa cells (2 X 10⁶ cell equivalents per IP) was subjected to chromatin immunoprecipitation using 4 μg of either a negative control antibody or Anti-Monomethyl-Histone H4
(Lys20) antibody and the Magna ChIP A Kit (Cat. #17-610). Successful immunoprecipitation of monomethyl-histone H4 (Lys20)-associated DNA fragments was verified by qPCR using GAPDH coding region ChIP Primers versus Control Primers corresponding to the GAPDH promoter.
Please refer to the EZ-Magna ChIP A (Cat. # 17-408) or EZ-ChIP (Cat. # 17-371) protocol for experimental details.

Dot Blot Analysis : Absurance Histone H3 Antibody Specificity Array (Cat. No. 16-667) and Absurance Histone H2A, H2B, H4 Antibody Specificity Array (Cat. No. 16-665), which contain histone peptides with various modifications were probed with Cat. No. 07-1570-S Anti-monomethyl Histone H4 (Lys20) at 1:1000 dilution. Proteins were visualized using a Donkey anti-rabbit IgG conjugated to HRP and a chemiluminescence detection system.

Western Blot Analysis:
0.1 μg/mL of proteins were visualized using a goat anti-rabbit secondary antibody conjugated to HRP and a chemiluminescence detection system (Please see figures)

Peptide Blocking Assay:
40 μg of histone H4 peptide containing monomethyl Lys20 abolished detection of histone H4 by anti-monomethyl-Histone H4 (Lys20) in immunoblot analysis of acid extracts of HeLa cells (Please see figures).

Research Category
Epigenetics & Nuclear Function

Research Sub Category
Chromatin Biology

Histones

Based on sequence homology, broad species cross-reactivity is expected

Recognizes monomethylated Histone H4
(Lys20), Mr 11 kDa.

Affinity Chromatography

Purified from serum by affinity chromatography. Supplied at 1 mg/ml in 0.1M Tris-Glycine (pH7.4), 15mM NaCl, 0.05% NaN3.

Stable for 1 year at -20°C from date of receipt.
Handling Recommendations: Upon receipt, and prior to removing the cap, centrifuge the vial and gently mix the solution. Aliquot into microcentrifuge tubes and store at -20°C. Avoid repeated freeze/thaw cycles, which may damage IgG and affect product performance.

Control
HeLa cell acid extracts.

Routinely evaluated by western blot analysis

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