Work

Analysis of Histone Antibody Specificity with Peptide Microarrays

Year: 2017

Type: article

Abstract: Post-translational modifications (PTMs) on histone proteins are widely studied for their roles in regulating chromatin structure and gene expression. The mass production and distribution of antibodies specific to histone PTMs has greatly facilitated research on these marks. As histone PTM antibodies are key reagents for many chromatin biochemistry applications, rigorous analysis of antibody specificity is necessary for accurate data interpretation and continued progress in the field. This protocol describes an integrated pipeline for the design, fabrication and use of peptide microarrays for profiling the specificity of histone antibodies. The design and analysis aspects of this procedure are facilitated by ArrayNinja, an open-source and interactive software package we recently developed to streamline the customization of microarray print formats. This pipeline has been used to screen a large number of commercially available and widely used histone PTM antibodies, and data generated from these experiments are freely available through an online and expanding Histone Antibody Specificity Database. Beyond histones, the general methodology described herein can be applied broadly to the analysis of PTM-specific antibodies.

Source: Journal of Visualized Experiments

DOI: 10.3791/55912

Authors: Evan M. Cornett, Bradley M. Dickson, Scott B. Rothbart

Institution: Van Andel Institute

Language: English

Topic: Genomics and Chromatin Dynamics

Subfield: Molecular Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: green

Funders: Van Andel Research Institute, National Institutes of Health, National Cancer Institute

Awards: Mechanisms Regulating DNA Methylation Maintenance in Chromatin, Mechanisms Regulating DNA Methylation Maintenance in Chromatin