Histone Arginine Citrullination Analysis

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Histone citrullination serves as a critical nexus linking innate immunity with epigenetic regulation, playing a central role in the pathogenesis of autoimmune diseases. CD BioSciences provides professional histone arginine citrullination analysis services, offering diverse detection solutions ranging from antibody-based assays to mass spectrometry, comprehensively covering the identification and quantitative analysis of low-abundance citrullination modifications.

Introduction to Histone Arginine Citrullination

Histone arginine citrullination, also known as deimination, is an irreversible enzymatic process catalyzed by peptidylarginine deiminases (PADs). It involves the conversion of a positively charged arginine residue into a neutral citrulline, resulting in the loss of a positive charge and a subtle mass increase of +0.984 Da. This modification plays crucial biological roles by altering chromatin structure, loosening histone-DNA interactions, and influencing gene expression. It is a key driver in immune-related processes, most notably in the formation of neutrophil extracellular traps (NETosis) during innate immune responses.

Dysregulation of histone citrullination is strongly linked to the pathogenesis of autoimmune diseases such as rheumatoid arthritis (RA) and multiple sclerosis (MS), as citrullinated neo-epitopes can become targets for autoantibodies, highlighting its significance as a major link between epigenetic regulation and immune dysfunction.

Citrullination catalyzed by PAD enzymes.

Fig.1 PAD-mediated citrullination. (Zhu D, et al., 2021)

Our Services

Building on our deep expertise in epigenetic and immune regulation, CD BioSciences delivers comprehensive histone arginine citrullination analysis services by integrating highly specific antibody-based enrichment with cutting-edge mass spectrometry. Our service is designed to overcome the significant challenges of studying this low-abundance and irreversible modification, providing you with robust and validated data to explore its dynamics in health and disease.

Customized Solution for Histone Arginine Citrullination Analysis

Global Citrullinome Profiling: Comprehensive identification and mapping of citrullination sites across core histones.
Site-Specific Targeted Quantification: Precise relative or absolute quantification of specific citrullination events (e.g., H3R2/R8/R17/R26cit).
PAD Enzyme Activity & Inhibition Studies: Analysis of citrullination changes in response to PAD activation, specific stimuli (e.g., calcium influx), or inhibitor treatment.
NETosis & Immune Response Analysis: Focused profiling of citrullination during neutrophil activation and extracellular trap formation.
Disease Mechanism Studies: Comparative analysis of citrullination profiles in autoimmune disease models or patient-derived samples.

Workflow of Histone Arginine Citrullination Analysis Service

Our specialized workflow is meticulously designed to ensure the accurate capture and analysis of the challenging histone arginine citrullination modification, providing reliable and biologically meaningful data for your research in immunology and disease mechanisms.

Project Design & Model Optimization

We begin with a collaborative consultation to design the optimal experimental strategy. This includes selecting the most relevant disease models (e.g., autoimmune, inflammatory) and defining precise stimulation protocols to effectively induce or modulate PAD enzyme activity for robust citrullination signal detection.

Pre-Analysis Sample Processing

To preserve the native citrullination state, we implement a calcium-stabilized histone extraction protocol. This critical step minimizes artifactual, non-enzymatic deimination during sample preparation, ensuring that the measured signal accurately reflects the in vivo biological condition.

Specific Enrichment Strategy

We employ a high-efficiency, multi-antibody enrichment strategy to selectively isolate citrullinated histones or peptides. This approach significantly enhances the target signal from the complex background, which is essential for analyzing this low-abundance modification.

High-Fidelity Mass Spectrometry Analysis

Enriched samples are analyzed using high-resolution mass spectrometry, specifically utilizing the parallel reaction monitoring (PRM) method on platforms like the Orbitrap Fusion™. This targeted approach ensures high sensitivity, accurate quantification, and confident identification of citrullination sites despite the small associated mass shift.

Rigorous Data Verification

All identifications and quantitative results are stringently validated using synthetic, isotopically labeled peptide standards. This step confirms the accuracy of site assignment and provides a solid foundation for reliable quantitative comparisons across samples.

Integrated Reporting & Biological Correlation

You receive a comprehensive report that not only presents the raw and processed data but also integrates the citrullination profile analysis. We correlate modification abundance with relevant disease scores or clinical parameters, delivering actionable insights that connect epigenetic changes to phenotypic outcomes.

Supported Sample Types

We support a diverse range of sample types, with specialized expertise in processing immunology and clinically relevant specimens for citrullination analysis.

Sample Types	Description
Immune Cells	Primary immune cells including neutrophils, macrophages, monocytes, T-cells, and B-cells, processed under resting, activated (e.g., with PMA/Iono, LPS), or NETosis-induced conditions.
Mammalian Cell Lines	Standard and engineered cell lines, particularly hematopoietic, synovial, or epithelial lines used in inflammation or autoimmune research.
Primary Tissues	Fresh or snap-frozen tissues such as synovium, skin, lymph nodes, and brain tissue.
Pre-Extracted Histones	Acid-extracted or affinity-purified histone samples from any biological source.
Clinical Specimens	Samples from autoimmune disease studies, including synovial fluid, serum, plasma, and tissue biopsies (e.g., from rheumatoid arthritis or psoriasis patients).
Archival FFPE Tissues	Formalin-fixed, paraffin-embedded tissue blocks or sections, accepted subject to a prior feasibility assessment for citrullination preservation.

*Optimal sample integrity is achieved through snap-freezing and dry ice shipment. We strongly recommend consulting our technical team before submission to confirm specific sample requirements, minimum amounts, and the optimal protocol for your research model.

Our Advantages

High-Specificity Enrichment Expertise: Use of rigorously validated antibodies and protocols for efficient capture of citrullinated histones, minimizing background.
High-Confidence Detection of a Subtle Modification: Optimized MS methods and data analysis pipelines to reliably identify the small +0.984 Da mass shift.
Focus on Immune & Disease Biology: Specialized support and experimental design for research in autoimmunity, NETosis, and inflammatory signaling.
End-to-End Scientific Partnership: Guidance from experimental design through complex data interpretation by specialists in immunology and epigenetics.

Beyond histone citrullination, CD BioSciences is your partner for comprehensive epigenetic analysis. We offer extensive services for studying methylation, acetylation, phosphorylation, ubiquitination, SUMOylation, formylation, succinylation, crotonylation and lactylation, among others. Our flexible platform allows for the creation of fully customized, multi-PTM profiling solutions tailored to your specific research questions in immunology, cancer biology, and beyond. Contact us to design an integrated strategy for your next project.

Reference

1. Zhu D, Zhang Y, Wang S. Histone citrullination: a new target for tumors[J]. Molecular Cancer, 2021, 20(1): 90.