Fluorescence in situ hybridization in combination with the comet assay and micronucleus test in genetic toxicology

Abstract

Comet assay and micronucleus (MN) test are widely applied in genotoxicity testing and biomonitoring. While comet assay permits to measure direct DNA-strand breaking capacity of a tested agent MN test allows estimating the induced amount of chromosome and/or genome mutations. The potential of these two methods can be enhanced by the combination with fluorescence in situ hybridization (FISH) techniques. FISH plus comet assay allows the recognition of targets of DNA damage and repairing directly. FISH combined with MN test is able to characterize the occurrence of different chromosomes in MN and to identify potential chromosomal targets of mutagenic substances. Thus, combination of FISH with the comet assay or MN test proved to be promising techniques for evaluation of the distribution of DNA and chromosome damage in the entire genome of individual cells. FISH technique also permits to study comet and MN formation, necessary for correct application of these methods. This paper reviews the relevant literature on advantages and limitations of Comet-FISH and MN-FISH assays application in genetic toxicology.

Figure 1

Example of SYBR-green-stained comet image from BLM-treated human leukocytes with telomeric PNA probes indicating the location of telomeric repeat sequences.

Figure 2

Example of DAPI-stained binucleated cell image from MMC-treated human leukocytes with centromeric (A) and whole chromosome painting (B) probes for chromosomes 7, 18 and X. MN (a) contains centromeric signal from chromosome X in (A) and whole chromosome probe signals from chromosome X in (B). MN (b) did not provide neither centromeric nor wcp signals.