New concepts to improve resolution and sensitivity of molecular cytogenetic diagnostics by multicolor fluorescence in situ hybridization.

BACKGROUND Routine application of multicolor fluorescence in situ hybridization (M-FISH) technology for molecular cytogenetic diagnostics has been hampered by several technical limitations. First, when using chromosome-specific painting probes, there is a limit in cytogenetic resolution of approximately 2-3 Mb, which can mask hidden structural abnormalities that have a significant clinical effect. Second, using whole chromosome painting probes, intrachromosomal rearrangements cannot be detected and the exact localization of breakpoints is often not possible. METHODS We suggest the use of multiplex-labeled region or locus- specific probes in combination with an optimal probe design to improve the sensitivity and resolution of the M-FISH technology. To allow the application of this assay in routine diagnostics, we developed a multipurpose image analysis system. RESULTS goldFISH was applied to the study of cryptic translocations in mental retardation patients and to the study of high-resolution breakpoint mapping in non-small cell lung cancer patients. For an individual with mental retardation, who had an apparently normal karyotype by G-banding, we detected an unbalanced translocation involving chromosomes 2 and 7. CONCLUSIONS In combination with optimally designed probe kits, goldFISH overcomes most of the present limitations of the M-FISH technology and results in virtually 100% reliability for detecting interchromosomal and intrachromosomal rearrangements.