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• W2045176586 abstract "OBJECTIVE: Determine the maximum number of chromosomes that can be detected using Oligo-FISH on the same slide in a single day. DESIGN: Single fluor labeled probes can be used to enumerate human chromosomes in interphase cells. Because of high stringency required in conventional FISH experiments, no more than 3 hybridizations can be performed sequentially. Because 4-6 hours are required per hybridization, it is impossible to perform more than one per day. Oligo-FISH probes are currently available in 5 fluors permitting enumeration of 5 chromosomes per hybridization. In addition, because of low stringency conditions and short hybridization time (5min), up to 6 successive FISH experiments can be performed on the same slide without significant signal loss. Performing FISH 3 times with a combination of 5 chromosomes (FISH 1: X, Yq12, 15, 17, 20; FISH 2: 3, 7, 8, 10, 16; FISH 3: 2, 4, 6, 9q12, 12) and 1 hybridization with 3 chromosomes (FISH 4: 1q12, 11, 18), 18 chromosomes in the same cell can be enumerated in record time. MATERIALS AND METHODS: Cytogenetic Slides from 5 chromosomally normal males were prepared by harvesting peripheral blood as previously described. ODN sequence Design: ODNs were selected in the regions where the satellite 2-3 pentamer or the α-satellite consensus sequence was underrepresented. The sequences were compared to the human whole genome database (NCBI) using BLAST. ODN probes have been designed to detect 18 human chromosomes that harbor specific repeats. FISH was performed as previously described. Image Acquisition was performed using a Nikon microscope equipped with a CCD camera. Images were captured using NIS elements software. Analytical Specificity and Sensitivity were calculated according to the Standards and Guidelines for Clinical Genetic Laboratories of the American College of Medical Genetics.S/N was calculated on 50 interphase nuclei. Images were segmented into signal and noise regions. Dividing the signal level to the noise level the ratio was obtained. RESULTS: We designed specific ODNs for 18 chromosomes. After each hybridization, all probes exhibited S/N>2 and sensitivity >98%. We successfully detected 18 chromosomes in 4 successive hybridizations in a single day. CONCLUSIONS: ODN probes' short length allows rapid hybridization and much less damage to cells, which are significant advantages for time critical procedures such as enumeration of chromosomes in interphase nuclei for PGD." @default.
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• W2045176586 date "2008-09-01" @default.
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• W2045176586 title "Oligonucleotide (ODN) fluorescence in situ hybridization (Oligo-FISH) allows enumeration of 18 chromosomes in 4 successive hybridizations performed in a single day" @default.
• W2045176586 doi "https://doi.org/10.1016/j.fertnstert.2008.07.441" @default.
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