Karyotyping human chromosomes by combinatorial multi-fluor FISH

Abstract

We have developed epifluorescence filter sets and computer software for the detection and discrimination of 27 different DNA probes hybridized simultaneously. For karyotype analysis, a pool of human chromosome painting probes, each labelled with a different fluor combination, was hybridized to metaphase chromosomes prepared from normal cells, clinical specimens, and neoplastic cell lines. Both simple and complex chromosomal rearrangements could be detected rapidly and unequivocally; many of the more complex chromosomal abnormalities could not be delineated by conventional cytogenetic banding techniques. Our data suggest that multiplex-fluorescence in situ hybridization (M-FISH) could have wide clinical utility and complement standard cytogenetics, particularly for the characterization of complex karyotypes.

Chromosomal alterations in small cell lung cancer revealed by multicolour fluorescence in situ hybridization

“…10,11 Before hybridization, metaphase preparations were enzymatically pretreated to remove cytoplasmic proteins and RNA molecules and to enhance the penetration of the probe. Exact pretreatment conditions varied between metaphase preparations depending on the volume of residual cytoplasm.…”

“…9 Multicolour-fluorescent in situ hybridization (M-FISH) is a recently developed technique designed to complement classical cytogenetic banding analysis. 10,11 Metaphase chromosomes prepared from fresh tumours subjected to short term culture are 'painted' with a combination of whole chromosome paints that allow each chromosome to be visualised in a unique colour (colour karyotyping). This technique, along with the technically similar spectral karyotyping (SKY), has greatly facilitated the identification of chromosome material involved in chromosomal rearrangements even in sup-optimal chromosome preparations.…”

American College of Medical Genetics guideline on the cytogenetic evaluation of the individual with developmental delay or mental retardation

“…In addition, techniques have been developed to differentially label each human chromosome with a unique color combination to obtain 24-color karyotypes 35–38. This allows the detection of structural chromosomal aberrations at a level of resolution similar to that of a routine karyotype (400 to 550 bands).…”

New Tools in Molecular Pathology

“…Because loss or gain of gene functions by point mutations or chromosome aberrations are crucial for the etiology and development of tumors, recurrence of changes in one locus serves as an indicator of genes playing a pathogenic role. Approaches for the genome-wide screening of such alterations include means of cytogenetics such as chromosome banding, 1 comparative genomic hybridization (CGH), 2,3 and 24-color chromosome painting by multicolor fluorescence in situ hybridization (FISH), 4,5 as well as means of molecular genetics such as loss of heterozygosity analysis. Thus, the search for new factors is based on a gene's location rather than on its known or assumed function.…”