MonoMethyl-Histone H4-K5 Rabbit pAb (P62805) for IF/ICC and ELISA

Core Product Specifications and Parameters

Biological Background: Histone H4 Function and Localization

• Function: Histone H4 is a core component of the nucleosome, wrapping and compacting DNA into chromatin. This limits DNA accessibility and plays a central role in transcription regulation, DNA repair, DNA replication, and chromosomal stability. Nucleosomes consist of two copies each of histones H2A, H2B, H3, and H4, forming an octamer around which 147 base pairs of DNA are wrapped. The histone code, mediated by post-translational modifications, regulates DNA accessibility and nucleosome remodeling. Related references: UniProtKB P62805

• Subcellular Localization: Histone H4 is primarily localized to the nucleus and associates with chromosomes. Acetylation can influence its subnuclear distribution; for instance, acetylated H4 localizes to the nucleus in step 11 spermatids. Related references: UniProtKB P62805

• Post-Translational Modifications: Histone H4 undergoes extensive post-translational modifications, especially on its N-terminal tail. Acetylation occurs at multiple lysines (e.g., K6, K9, K13, K17, K32, K92; see PMID:19608861). Methylation is found at arginine 4 (asymmetric dimethylation by PRMT1; PMID:11387442) and lysine 21 (monomethylation; PMID:12086618). Monomethylation at lysine 5 (K5me1) is a specific epigenetic mark recognized by various reader proteins and implicated in transcriptional regulation. Other modifications include phosphorylation, citrullination, and ubiquitination, collectively forming a complex regulatory code. Related references: PMID:19608861, PMID:11387442, PMID:12086618

Experimental Guidance and Technical Tips

• Immunogen Design: This antibody was raised against a synthetic peptide corresponding to the N-terminal region of human Histone H4 monomethylated at K5. When designing similar antibodies, ensure the peptide is highly pure and the modification is confirmed by mass spectrometry. Consider using a non-methylated peptide as a negative control in your assays to verify specificity.
• IF/ICC: For immunofluorescence, standard fixation with 4% paraformaldehyde and permeabilization with 0.1-0.5% Triton X-100 is typically suitable. Block non-specific binding with 5% BSA or serum. Epitope accessibility may vary; optimizing antigen retrieval could be considered. Since histone proteins are nuclear, expect nuclear staining. Include no-primary antibody controls to assess background.
• ELISA: For coating, dilute the antibody or antigen in carbonate/bicarbonate buffer (pH 9.6). Titrate antibody concentration to determine optimal signal-to-noise ratio. Use monomethylated peptide as positive control and unmodified peptide as negative control to confirm specificity.
• General: As with any modification-specific antibody, lot-to-lot variation may occur; validate new lots in your specific experimental system. Cross-reactivity is predicted for human, mouse, and rat based on sequence conservation; test on your species of interest.

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