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Eils, R; Uhrig, S; Saracoglu, K; Sätzler, K; Bolzer, A; Petersen, I; Chassery, J; Ganser, M; Speicher, MR.
An optimized, fully automated system for fast and accurate identification of chromosomal rearrangements by multiplex-FISH (M-FISH).
Cytogenet Cell Genet. 1998; 82(3-4):160-171 Doi: 10.1159/000015092
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Leading authors Med Uni Graz: Speicher Michael
Co-authors Med Uni Graz: Uhrig Sabine
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Abstract:: Multiplex-FISH (M-FISH) is a recently developed technique by which each of the two dozen human chromosomes-the 22 autosomes and the X and Y sex chromosomes-can be stained or "painted" with uniquely distinctive colors. Using a combinatorial labeling technique and a specially designed filter set, each DNA probe can be identified by its unique spectral signature. Here we present several significant optimizations of the M-FISH technology. First, a new strategy for labeling the probes is described which allows for easy and fast production of the complex M-FISH probe mix. Second, a newly developed, completely motorized microscope equipped with an eight-position filter wheel and a new generation of filter sets is presented that allows fully automatic imaging of a complete metaphase spread within seconds. Third, to determine the characteristic spectral signatures for all different combinations of fluorochromes, we developed a novel multichannel image analysis method. The spectral analysis is solely guided by the image information itself and does not require any user interaction. A complete analysis of a metaphase spread can be accomplished in less than 3 min. Sophisticated built-in quality controls were developed, and the value of visual inspection of M-FISH images as a simple means of controlling the computer-generated chromosome classification are illustrated. In addition, we discuss advantages of adding new fluorochromes to the traditionally used five fluorochromes.
Find related publications in this database (using NLM MeSH Indexing): Chromosome Aberrations -; Chromosome Banding - methods; DNA, Neoplasm - analysis; Fluorescent Dyes - analysis; Gene Rearrangement - analysis; Humans - analysis; Image Processing, Computer-Assisted - methods; In Situ Hybridization, Fluorescence - methods; Indoles - methods; Microscopy, Fluorescence - methods