Detailed Genome-Wide Screening for Inter- and Intrachromosomal Abnormalities by Sequential G-Banding and RxFISH Color Banding of the Same Metaphase Cells

Abstract

While the now-classic chromosome banding methods, such as G-banding, remain the techniques of choice for the initial screening for karyotypic abnormalities, sometimes chromosomal rearrangements involve segments too small or too similarly banded to be detected or described adequately by these techniques. The necessity to use a genome-wide, fluorescence in situ hybridization (FISH)-based screening technique as a complement to G-banding is especially obvious in cases where the information obtained by the latter analysis does not provide an adequate guide to the choice of probes for chromosome-specific FISH. Furthermore, the same metaphase cells should ideally be used for both G-banding and FISH analysis to overcome the scarcity of metaphases observed in many cases and to ensure the correct interpretation of chromosomal aberrations in cytogenetically unstable neoplasms with massive cell-to-cell karyotypic variability. We describe a protocol which enables cross-species color banding (RxFISH), a new FISH-based screening technique that simultaneously imparts specific color banding patterns on all chromosomes, of preparations that have been G-banded and mounted for up to several years, as well as a procedure allowing chromosome-specific painting of the same metaphase cells to resolve whatever doubts persist after the preceding G-banding and RxFISH analyses. This approach makes possible a detailed, genome-wide screening for inter- and intrachromosomal abnormalities including archival cases whose karyotypic rearrangements had been incompletely identified by G-banding.