Image selection is critical in the dicentric chromosome assay in cytogenetic biodosimetry

The dicentric chromosome assay (DCA), recognized as the gold standard for biological dosimetry in emergency radiation exposure, involves multiple key steps. Among these, metaphase image selection remains ambiguous and particularly challenging for beginners. This study evaluates the impact of metaphase image selection on dicentric (Dic) frequency during initial training.

Peripheral blood (PB) from three healthy donors was irradiated with 2 Gy X-rays and incubated at 37 °C for 2 h for DNA repair. PB samples were then cultured for 48 h with phytohemagglutinin and colcemid. After fixation, metaphase spreads were prepared, Giemsa-stained, and images were captured by specific microscope imaging and image processing software. Three beginners independently selected and analyzed 50 metaphase images each. A trainer with experience in performing radiation emergency related DCA then reviewed and removed unsuitable images. The beginners reanalyzed Dic frequencies, supplementing with additional images as needed to reach 50 metaphases. Dic frequencies were compared before and after image refinement.

Dic frequencies were lower in metaphase image pools selected by beginners compared to those refined by the trainer. The novices' selections included numerous over-condensed metaphases, making Dic detection difficult.

Blood culture for 48 h permanent colcemid treatment increases the occurrence of over-condensed chromosomes, affecting Dic scoring. This study highlights the importance of proper training in metaphase image selection and structured education on quality assessment, including the use of specific microscope imaging and image processing software to ensure accurate biological dosimetry.