title: Automation of the cytokinesis-block micronucleus cytome assay by laser scanning cytometry and its potential application in radiation biodosimetry.
abstract: Here we describe the adaptation of laser scanning cytometry (LSC) to measure micronuclei (MN) automatically in lymphocytes. MN frequencies were determined in irradiated human lymphocytes using the cytokinesis-block technique, and the results from LSC were compared with visual scoring results obtained from slides of cells stained using Fast Green and 4',6-diamidino-2-phenylindole (DAPI). This fluorescent approach allowed clear identification of binucleated cells and detection of MN. The dose responses measured visually and by LSC showed similar trends and correlated positively (r = 0.9689; P < 0.0001). High-content analysis was developed to further automatically score MN within mono-, tri- and tetra-nucleated cells and to determine the nuclear division index and nuclear circularity values. The high-throughput nature of LSC can provide unique advantages in future DNA damage diagnostics in experimental and epidemiological studies. Importantly, it allows for co-detection of other biomarkers of interest within a single lymphocyte, and further development of this capability is anticipated.
authors.affiliation: CSIRO Food and Nutrition Flagship, Nutrigenomics and DNA damage, Adelaide, Australia.
authors.forename: Maxime
authors.initials: M
authors.lastname: François
authors.affiliation: CSIRO Food and Nutrition Flagship, Nutrigenomics and DNA damage, Adelaide, Australia; Section of Investigative Medicine, Department of Medicine, Imperial College London, London, United Kingdom.
authors.forename: Kevin
authors.initials: K
authors.lastname: Hochstenbach
authors.affiliation: CSIRO Food and Nutrition Flagship, Nutrigenomics and DNA damage, Adelaide, Australia.
authors.forename: Wayne
authors.initials: W
authors.lastname: Leifert
authors.affiliation: CSIRO Food and Nutrition Flagship, Nutrigenomics and DNA damage, Adelaide, Australia.
authors.forename: Michael Felix
authors.initials: MF
authors.lastname: Fenech
mesh_terms: D057205:Automation, Laboratory; D048749:Cytokinesis; D006801:Humans; D007211:Indoles / Q000737:chemistry; D047569:Laser Scanning Cytometry / Q000379:methods*; D008214:Lymphocytes / Q000166:cytology / Q000528:radiation effects; D015162:Micronucleus Tests / Q000379:methods*; D014965:X-Rays
chemical_list: D007211:Indoles; C007293:DAPI
keywords: Micronucleus assay; automation; laser scanning cytometry