Chromosomics: Detection of Numerical and Structural Alterations in All 24 Human Chromosomes Simultaneously Using a Novel OctoChrome FISH Assay

Abstract

Fluorescence in situ hybridization (FISH) is a technique that allows specific DNA sequences to be detected on metaphase or interphase chromosomes in cell nuclei 1 . The technique uses DNA probes with unique sequences that hybridize to whole chromosomes or specific chromosomal regions, and serves as a powerful adjunct to classic cytogenetics. For instance, many earlier studies reported the frequent detection of increased chromosome aberrations in leukemia patients related with benzene exposure, benzene-poisoning patients, and healthy workers exposed to benzene, using classic cytogenetic analysis 2 . Using FISH, leukemia-specific chromosomal alterations have been observed to be elevated in apparently healthy workers exposed to benzene [3][4][5][6] , indicating the critical roles of cytogentic changes in benzene-induced leukemogenesis.Generally, a single FISH assay examines only one or a few whole chromosomes or specific loci per slide, so multiple hybridizations need to be conducted on multiple slides to cover all of the human chromosomes. Spectral karyotyping (SKY) allows visualization of the whole genome simultaneously, but the requirement for special software and equipment limits its application 7 . Here, we describe a novel FISH assay, OctoChrome-FISH, which can be applied for Chromosomics, which we define here as the simultaneous analysis of all 24 human chromosomes on one slide in human studies, such as chromosome-wide aneuploidy study (CWAS) 8 . The basis of the method, marketed by Cytocell as the Chromoprobe Multiprobe System, is an OctoChrome device that is divided into 8 squares, each of which carries three different whole chromosome painting probes (Figure 1). Each of the three probes is directly labeled with a different colored fluorophore, green (FITC), red (Texas Red), and blue (Coumarin). The arrangement of chromosome combinations on the OctoChrome device has been designed to facilitate the identification of the non-random structural chromosome alterations (translocations) found in the most common leukemias and lymphomas, for instance t(9;22), t(15;17), t(8;21), t(14;18) 9 . Moreover, numerical changes (aneuploidy) in chromosomes can be detected concurrently. The corresponding template slide is also divided into 8 squares onto which metaphase spreads are bound (Figure 2), and is positioned over the OctoChrome device. The probes and target DNA are denatured at high-temperature and hybridized in a humid chamber, and then all 24 human chromosomes can be visualized simultaneously.OctoChrome FISH is a promising technique for the clinical diagnosis of leukemia and lymphoma and for detection of aneuploidies in all chromosomes. We have applied this new Chromosomic approach in a CWAS study of benzene-exposed Chinese workers 8,10 .

Persistence of Breakage in Specific Chromosome Bands 6 Years after Acute Exposure to Oil

“…All these chronic exposure studies were performed using G banding analysis, and the same analysis was employed in the present study. The loss of chromosomes 5 and 7, deletions of 5q31 and 7q22, and translocations of t(9;22), t(15;17), t(8;21) and t(14;18) have also been detected using the FISH technique [ 37 – 40 ].…”

Toxicological Risk Assessment for Beginners

“…Recent investigations continue to assess and report the frequent detection of increased chromosomal aberrations among benzene exposed leukemia patients (Paz-y-Mino et al 2008;Kim et al 2010;Zhang et al 2011;Ji and Zhang 2012;Zhang et al 2012). For additional studies and investigations on the cytogenetic effects of benzene, please see Kim et al (2004), Holeckova et al (2004), Zhang et al (2002), Marcon et al (1999), Smith et al (1998), Zhang et al (1996), Ciccone et al (1993), Katz et al (1992), Shannon et al (1992), Cuneo et al (1992), Robertson et al (1991), Garson (1984), Rowley (1983) and Mitelman et al (1981).…”

Formaldehyde, Hematotoxicity, and Chromosomal Changes—Response

“…Our study methods called for counting all scorable metaphases on a subject's slide with a minimum of 150 cells counted per study subject (2,3). Ã A reference for the method, called OctoChrome FISH, was provided in reference 45 (10) of that publication (2), and additional details and references were provided in the follow-up article in Carcinogenesis (3,(10)(11)(12). In brief, we used an automated metaphase finder to detect metaphases on each subject's slide.…”