A CRITICAL-EVALUATION OF CENTROMERIC LABELING TO DISTINGUISH MICRONUCLEI INDUCED BY CHROMOSOMAL LOSS AND BREAKAGE IN-VITRO

The in vitro micronucleus assay in conjunction with CREST-staining and fluorescence in situ hybridization (FISH) with centromere-specific DN A probes is being increasingly utilized for the detection of clastogen ic and aneuploidy-inducing agents. Although potentially powerful techn iques, both methods have unique characteristics that can influence sam ple processing and the interpretation of results. In this article, the use of the CREST and the FISH modifications of the in vitro micronucl eus assay have been used to characterize the origin of the micronuclei induced by cyclophosphamide, 4,4'-methylene-bis(2-chloroaniline), 4-n itroquinoline N-oxide and ionizing radiation in metabolically competen t MCL-5 cells or a derived cell line lacking metabolic activation. Usi ng these results and our previous experiences with these techniques, a detailed comparison including the strengths and limitations of each t echnique as well as potential problems in performing each assay and in analyzing the data is discussed, In spite of their limitations, our r esults to date indicate that CREST-staining as well as FISH with centr omere-specific DNA probes can be used to accurately distinguish micron uclei formed from chromosome loss from those originating from chromoso me breakage and that these techniques can be valuable complements to t he in vitro micronucleus assay. (C) 1997 Elsevier Science B.V.