Use of centromeric probe to identify micronuclei origin and its advantages in genetic toxicology studies

Fluorescence in situ hybridization (FISH) and karyotyping have long been considered essential for chromosomal examination to analyse the stable chromosome aberrations such as translocations after exposure to genotoxic agents. To improve speed of chromosomal analysis in genetic toxicology investigations, we employed the cytokinesis-block micronucleus (CBMN) assay with human pan-centromeric probes, to measure chromosome segregation errors and to differentiate mechanism of action of genotoxic agents. Whole-blood collected from healthy volunteers were exposed to X-rays, bleomycin (BLM), Colchicine (COL) and Mitomycin-C (MMC), arrested at cytokinesis stage and processed for conventional giemsa staining and pan-centromeric FISH to analyse the composition of MN to differentiate between aneugen and clastogen effects. One-way ANOVA with Tukey's multiple comparison test exhibited a significant (p < 0.001) increase in the MN observed using giemsa stained and centromere FISH binucleated cells in all those genotoxic agents when compared to its unexposed lymphocytes cultures. The CBMN with centromere FISH, predominantly shows MNCN^-ve cells in lymphocytes exposed to X-rays (67.4 % and 83.3 %), BLM (79.4 % and 79.2 %) and MMC (81.5 % and 73.2) while, COL treatment resulted in significantly (p < 0.05) higher MNCN^+ve cells (79.6 % and 82.4 %) for the different concentrations of those agents. Centromere FISH enables to identify the origin of MN induced by different genotoxic agents. Thus, a significant development in genetic toxicology is heralded by the incorporation of centromeric probe in modern analytical workflows.