Measurement and characterization of micronuclei in exfoliated XXXX cells by fluorescence in situ hybridization with a centromeric probe.

@entity1 @entity4523 @entity74 @entity2667
The @entity4523 ( @entity4523 ) assay in @entity1 exfoliated cells has been widely used to detect the genotoxic effects of environmental mutagens, infectious agents and @entity74 . Substantial variability characterizes the @entity4523 frequencies reported by different research groups. One reason for this may be the restricted resolution power of the Feulgen-Fast-Green staining that is routinely used. Here we describe a new version of the @entity4523 assay that employs fluorescent @entity2667 staining along with fluorescence in situ hybridization (FISH) with a centromeric probe. Buccal and urothelial cells were collected from 5 healthy unexposed female volunteers and 55,000 cells analyzed for @entity4523 frequency and abnormal nuclear events. The Feulgen-Fast-Green and the new fluorescent staining produced very similar results. The frequency of @entity4523 in buccal cells was 0.145 +/- 0.118% and in urothelial cells 0.083 +/- 0.074%. No correlation was found between the frequencies of @entity4523 in the two types of exfoliated cells. FISH with a centromeric probe allowed @entity4523 containing whole chromosomes with a centromere to be differentiated from those containing only acentric fragments. The former appear as a result of chromosome lagging in mitosis, while those without a centromere are due to chromosome breakage. In urothelial cells 43% of @entity4523 were centromere-negative and in buccal cells -44%. Fluorescent staining provided more accurate scoring of degenerative cells than standard Feulgen-Fast-Green staining. The combined frequency of pycnotic cells, "broken eggs" and cells with fragmented nuclei did not exceed 2%, while that of karyorrhexis and karyolysis together was as high as 21%. Significant interindividual variability was found in the frequency of cells with karyolysis and karyorrhexis. Thus, the new version of @entity4523 assay allows for @entity4523 to be scored more precisely, the mechanism of @entity4523 formation to be determined and abnormal nuclear events to be readily identified in exfoliated @entity1 cells. It is therefore ideal for studying genotoxicity in @entity1 populations using exfoliated cells from the mouth, bladder and nose.
