Validation of Chromosome Painting as a Biodosimeter in Human Peripheral Lymphocytes Following Acute Exposure to Ionizing Radiationin Vitro

Abstract

Fluorescence in situ hybridization with chromosome-specific composite DNA probes ('chromosome painting') appears to be a useful tool for quantifying symmetrical cytogenetic damage. However, a thorough comparison between chromosome painting and the conventional methods of GTG-banding and dicentric analysis has not been performed. We have undertaken the validation of chromosome painting using human blood exposed in vitro to 137Cs gamma-rays at doses ranging from 0 to 400 cGy, then cultured according to standard procedures and harvested at 52 h. For painting, bound probes were detected either with fluoresceinated avidin and counterstained with propidium iodide, or with ChromoBlue WCP Probe and Giemsa. The first approach utilizes ultraviolet excitation in which painted chromosomes appear yellow and the remaining chromosomes appear red. The ChromoBlue labelling approach requires ordinary light microscopy in which painted chromosomes appear dark blue and the remaining chromosomes appear light blue. With each method, exchanges between painted and unpainted chromosomes appear bi-coloured. Because only a fraction of all possible exchanges are detected, the number of metaphases examined is adjusted according to the fraction of the genome painted. We have performed painting by several methods, including fluorescence with chromosome 4 probe alone, fluorescence with probes for chromosomes 1, 3 and 4 simultaneously, and chromogenic painting with chromosome 4 probe alone. The results obtained by the various painting methods were compared with GTG-banded cells which were examined for both translocations and dicentrics. In addition, unbanded metaphases stained with Giemsa were scored for dicentrics. Our data show that the frequency of chromosome exchanges detected by painting and banding agree with each other and with the number of dicentrics seen in unbanded cells, at least at doses of < or = 200 cGy.

Preferential reduction of dicentrics in reciprocal exchanges due to the combination of the size of broken chromosome segments by radiation

“…However, the actual observed rates of radiation-induced translocations are higher in general than that of dicentrics (1.63-9 fold, Lucas et al 1989;1-12.33 fold, Cremer el al. 1990; 1.5-2 fold, Natarajan et al 1992; 1-3 fold, Tucker et al 1993;1.67-4 fold, Boei et al 1996; greater than 1, VirsikPeuckert et al 1997;1.4 fold, Finnon et al 1995;1.3-3.75 fold, Luomahaara et al 1999;1.55-2.79 fold, Schmid et al 1992;1.11-3.67 fold, Bauchinger et al 1993; an observed high number in translocations at most dose points, Nakano et al 1993;1.2 fold, Knehr et al 1999). A study previously reported from our laboratory (Kanda et al 1996), in which cells in the second division and misclassification of aberrations were carefully excluded, also showed a slightly higher number of translocations, although the difference was not statistically significant.…”

Chromosome Painting in Biological Dosimetry: Assessment of the Ability to Score Stable Chromosome Aberrations Using Different Pairs of Paint Probes

“…In an in vitro study using ionizing radiation exposure with 137Cs gamma rays, Tucker et al (9) compared translocation detection between G-banded chromosomes and painting of chromosome 4 and chromosomes 1, …”

Second cancers after radiotherapy: any evidence for radiation-induced genomic instability?

“…For example, chromosome aberrations are the standard biodosimeter for radiation exposure (Tucker et al, 1993;Bauchinger, 1998;Finnon et al, 1999). This role reflects the relative ease of detecting aberrations and well-defined dose-response relationships.…”