current challenges and limitations in antibody-based detection of citrullinated histones

Abstract

Studies on NETosis demand reliable and convenient markers to monitor the progress of this form of cell death. Because a determining step in the release of nuclear chromatin NETs requires the conversion of arginine residues to citrulline residues in histones by peptidylarginine deiminase, citrullinated histones can provide such a marker. Here, we evaluate antibody reagents for the detection of citrulline residues in histones and observe alarming differences between commercial antisera and mouse and rabbit monoclonal antibodies in their ability to detect their nominal target residues. Differences between antibodies that are currently used to detect citrulline residues in histones could jeopardize efforts to reach a scientific consensus and instead lead to inconsistent and even conflicting conclusions regarding the regulation of histone deimination. Our results will assist others in planning their initial or ongoing studies on peptidylarginine deiminase activity with the use of currently available antibodies. Further, we argue that, along with the careful attention to experimental conditions and calcium concentrations, validated antibody reagents are urgently needed to avoid possible setbacks in the research on NETosis.