Affi-BAMS™: A Robust Targeted Proteomics Microarray Platform to Measure Histone Post-Translational Modifications

Area of Science:

• Biochemistry
• Molecular Biology
• Epigenetics

Background:

• Accurate quantification of post-translational modifications (PTMs) is essential for biological and pharmacological research.
• Histone PTMs play a critical role in regulating chromatin structure and gene expression.

Purpose of the Study:

• To evaluate the Affi-BAMS™ platform for quantitative analysis of histone PTMs.
• To demonstrate the platform's capability in resolving complex PTMs on histones H3 and H4.

Main Methods:

• Utilized the Affi-BAMS™ (epitope-directed affinity bead capture/MALDI MS) platform.
• Employed H3 and H4 histone peptides and isotopically labeled derivatives for calibration.
• Analyzed nuclear cellular lysates and MCF7 cell line models.

Main Results:

• Achieved a dynamic range of >3 orders of magnitude with <5% technical precision (CV).
• Successfully resolved heterogeneous N-terminal histone PTMs from 100 µg of starting material.
• Demonstrated monitoring of dynamic histone H3 acetylation and methylation events, including SILAC quantification.

Conclusions:

• Affi-BAMS™ provides an efficient and effective approach for analyzing dynamic epigenetic histone marks.
• The platform's multiplexing capacity enhances its utility for PTM analysis.
• This method is critical for advancing studies on chromatin regulation and gene expression.