Characterization of cosmids and telomeres in cytogenetic preparations by 3D confocal fluorescence

OBJECTIVE: To analyze, with fluorescent probes, by three-dimensional (30) e mission patterns, fluorescence in situ hybridization (FISH) preparations (c osmids, telomeres) and to perform factor analysis of medical image sequence s (FAMIS); to use FISH to track relevant DNA sequences in cell nuclei durin g interphase and in mitotic chromosomes; and to use cytogenetic techniques, resulting in flat preparations of whole cells that are assumed to preserve probe access to their targets. STUDY DESIGN: The study design entailed labeling targets by probes (sequenc es labeled by fluorescein isothiocyanate) in nuclei and/or chromosomes stai ned by propidium iodine. Visualization of targets was improved when 30 sequ ences of images obtained on a single photomultiplier detector of the confoc al microscope by z stepping were investigated by FAMIS. RESULTS: Factors and factor images showed that targets could be detected an d differentiated in focal planes inside nuclei or chromosomes. CONCLUSION: It is possible to localize cosmids in cell n nuclei at interpha se and telomeres in mitotic chromosomes by means of 30 sequences of images.