A control for the day-to-day normalization of the flow cytometry γ-H2AX assay for clinical routine.

Abstract

The phosphorylation of histone H2AX (γ-H2AX) at the DNA double-strand break (DSB) site is frequently used for quantifying DSBs and may be useful as a biomarker for clinical applications. We have previously reported a flow cytometry-based quantification of γ-H2AX for clinical routine. One major challenge, however, is the lack of a control sample for normalization of the day-to-day variation of the flow cytometry γ-H2AX assay. Here, we report development of a mix-control sample containing peripheral blood mononuclear cells (PBMC) from 10 control individuals, for normalization of day-to-day variation of the flow cytometry-γ-H2AX assay. We showed that control individuals sampled on different days show an average day-to-day variation (CV) of 34%, which was reduced to 12% after normalization to the control sample. The normalization allowed detection of radiosensitivity of lymphoblastoid cell lines from ataxia telangiectasia patients, sampled over three days. The mix-control sample, consisting of 10 control individuals' PBMC, can be used as a control sample to normalize for day-to-day variation of the γ-H2AX assay. The use of this sample will facilitate integration of the γ-H2AX assay into clinical routine. © 2018 International Clinical Cytometry Society.