TriMethyl-Histone H4-K20 Rabbit Polyclonal Antibody for WB/IHC/IF (P62805)

Core Product Specifications and Parameters

Biological Background: Histone H4 Function and Localization

Histone H4 is one of the core histones that constitute the nucleosome, the fundamental repeating unit of chromatin. Along with histones H2A, H2B, and H3, it forms an octamer around which approximately 147 base pairs of DNA are wrapped, compacting the genome into the nucleus and regulating DNA accessibility. Histone H4 plays a central role in transcription regulation, DNA repair, DNA replication, and chromosomal stability. The dynamic modulation of chromatin structure is largely achieved through a complex set of post-translational modifications (PTMs) on histone tails, collectively termed the "histone code." Related references: UniProt P62805

Histone H4 is primarily localized in the nucleus and associates with chromosomes. Acetylated H4 is also found in the nucleus during specific stages of spermatogenesis (step 11 spermatids). Related references: UniProt P62805

The lysine 20 residue of histone H4 (H4K20) is subject to multiple methylation states, including mono-, di-, and trimethylation. Trimethylation of H4K20 (H4K20me3) is a hallmark of constitutive heterochromatin and is involved in gene silencing, DNA damage response, and cell cycle regulation. This modification is dynamically regulated by methyltransferases (e.g., SUV4-20H1/2) and demethylases. Numerous other PTMs on H4 have been identified by mass spectrometry and other techniques, highlighting the complexity of histone regulation. Related references: PMID:12086618, PMID:15964846, PMID:17967882

Experimental Guidance and Technical Tips

• The immunogen is a synthetic peptide corresponding to the trimethylated K20 region of human histone H4. This antibody is expected to recognize H4K20me3 specifically, but cross-reactivity with other H4 methylation states or unmodified H4 should be ruled out experimentally using appropriate peptide competition assays or knockout/knockdown models.
• For Western blot (WB) detection, histone-enriched nuclear extracts or acid-extracted histones are recommended as the sample type. Optimize the primary antibody concentration (e.g., start at 1:500–1:2000) and use a suitable secondary antibody conjugate.
• In immunohistochemistry (IHC-P), antigen retrieval with citrate buffer (pH 6.0) or EDTA buffer (pH 8.0) may improve epitope availability; optimize the retrieval method empirically.
• For immunofluorescence (IF/ICC), permeabilization with 0.1–0.5% Triton X-100 or digitonin is suggested. Fixation conditions (e.g., 4% PFA) may affect epitope accessibility; test different protocols.
• When using in ELISA or dot blot (DB) assays, consider using BSA-blocked plates and a biotinylated detection system for enhanced sensitivity.
• Always include a negative control (e.g., unmodified H4 peptide or IgG control) and validate antibody specificity in the relevant sample system using mass spectrometry or other orthogonal methods.
• The antibody shows predicted wide-range cross-reactivity (Human, Mouse, Rat, and others), but validation in each species of interest is advised.

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