A semi-automated micronucleus-centromere assay to assess low-dose radiation exposure in human lymphocytes

Abstract

Purpose: The in vitro micronucleus (MN) assay is a reliable method to assess radiation-induced chromosomal damage in human peripheral blood lymphocytes. It is used to evaluate in vivo radiation over-exposure and to assess in vitro chromosomal radiosensitivity. A limitation of the MN assay is the relatively high and variable spontaneous MN frequency that restricts low-dose estimation to doses of about 0.3 gray (Gy). As radiation-induced MN mainly contain acentric fragments and spontaneous MN originate from lagging chromosomes, both MN types can be distinguished from each other by using fluorescence in situ hybridisation (FISH) with a pan-centromeric probe. The aim of this study was to investigate if the sensitivity, reliability and processing time of the MN assay can be enhanced by combining the automated MN assay with pan-centromere scoring.

Materials and methods: Blood samples from 10 healthy donors were irradiated in vitro with low doses of gamma-rays. Dose response curves were determined for fully-automated and semi-automated MN scoring and semi-automated scoring of centromere negative MN (MNCM−).

Results: A good correlation was obtained between fully-automated and semi-automated MN scoring (r² = 0.9973) and between fully automated MN scoring and semi-automated scoring of MNCM− (r² = 0.998). With the Wilcoxon test, a significant p value was obtained between 0 and 0.2 Gy for the fully-automated MN analysis, between 0 and 0.1 Gy for semi-automated MN analysis and between 0 and 0.05 Gy for semi-automated scoring of MNCM−.

Conclusion: The semi-automated micronucleus-centromere assay combines high-speed MN analysis with a more accurate assessment in the low-dose range which makes it of special interest for large-scale radiation applications.

Keywords::

• Semi-automated micronucleus-centromere assay
• biological dosimetry
• fluorescent in situ hybridisation
• low-dose radiation exposure
• dose response curves

Acknowledgements

The authors wish to thank Prof. D. Van der Merwe for the assistance in performing the irradiations. We also thank all volunteers who participated in this study.

Declaration of interest

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of the paper. This report is work commissioned by the National Institute for Health Research. The views expressed in this publication are those of the author(s) and not necessarily those of the NHS, the National Institute for Health Research or the Department of Health. The work was supported by National Research Foundation – iThemba LABS (Laboratory for Accelerator Based Sciences), South Africa and a University Development Cooperation ‘VLIR Own Initiative Programme’ between Belgium and South Africa (ZEIN 2005PR309) and by a grant of the Research Foundation Flanders (FWO, No 1.5.080.08).