Scoring of radiation‐induced microuclei in cytokinesis‐blocked human lymphocytes by automated image analysis

Abstract

The micronucleus assay in human lymphocytes is, at present, frequently used to assess chromosomal damage caused by ionizing radiation or mutagens. Manual scoring of micronuclei (MN) by trained personnel is very time-consuming, tiring work, and the results depend on subjective interpretation of scoring criteria. More objective scoring can be accomplished only if the test can be automated. Furthermore, an automated system allows scoring of large numbers of cells, thereby increasing the statistical significance of the results. This is of special importance for screening programs for low doses of chromosome-damaging agents.In this paper, the first results of our effort to automate the micronucleus assay with an image-analysis system are represented. The method we used is described in detail, and the results are compared to those of other groups. Our system is able to detect 88% of the binucleated lymphocytes on the slides. The procedure consists of a fully automated localization of binucleated cells and counting of the MN within these cells, followed by a simple and fast manual operation in which the false positives are removed. Preliminary measurements for blood samples irradiated with a dose of 1 Gy X-rays indicate that the automated system can find 89% f 12% of the micronuclei within the binucleated cells compared to a manual screening.

Cytokinesis-Block Micronucleus Assay in Human Glioma Cells Exposed to Radiation

“…Automated scoring of MN by computerized image analysis systems is an interesting alternative to tedious manual scoring. It gives results comparable to those obtained by manual scoring (Castelain et al, 1993, Verhaegen et al, 1994 with even better reproducibility (Thierens et al, 1997). However, automated MN scoring is time-consuming and is thus applicable only to a limited number of slides (Thierens et al, 1997).…”

Cytokinesis-block micronucleus cytome assay

“…For this goal to be achieved, it is essential that scoring criteria are well developed and that a robust slide preparation protocol be put in place and that slide preparations be permanent so that they can be re-examined visually if necessary. Automated image analysis systems have recently been developed for scoring of MNi in mammalian cells [66][67][68][69][70][71] (see http://www.imstar.fr/applications/genotoxicity/micronucleiassay/; http://www.metasystems.de/products/metafer/ metafer_msearch.htm#Micronucleus; http://www.biocompare.com/techart.asp?id=1247) but these systems do not take into account other important events such as NPBs, NBUDs, necrosis, apoptosis and cytostasis, which are essential for the correct interpretation of the result obtained. In the future, we should expect to have an automated system that can score reliably the various end points possible with the CBMN Cyt assay outlined in this paper.…”

“…This provides the opportunity to test, using the same system, the extent to which the in vitro test is predictive of in vivo genotoxic events. The adoption of the CBMN Cyt assay by the pharmaceutical industry 57,58,63 , its application in human biomonitoring of genotoxic exposures 24,60,61 and its increasing application in preventive medicine and nutrition 62,64,65 [66][67][68][69][70][71] .…”

In vitro micronucleus assay scored by flow cytometry provides a comprehensive evaluation of cytogenetic damage and cytotoxicity

“…These methods are based on image analysis, laser scanning cytometry, or flow cytometry [10][11][12][13][14][15][16][17][18][19][20].…”