Multicolor FISH using a novel spectral bioimaging system

FISH (fluorescence in situ hybridization) is a cytogenetic technique for locating and quantifying small genetic defects that cannot be detected by traditional karyotyping, the banding analysis of stained chromosomes. The selectivity of FISH holds the promise for the accurate, low cost and fast diagnosis of genetic disorders, birth defects, and various types of cancer that are not detectable by other means. To aid investigators with their FISH measurements, filter-based digital imaging systems are available to enhance and analyze faint FISH images. The accuracy and reliability of traditional FISH measurements is generally compromised when there is (1) sample autofluorescence, (2) image movement, e.g., due to lack of registration between images acquired through different filter sets, or (3) spectral overlap between fluorescent DNA probe emission spectra. SpectraCube TM is an interferometric imaging method which is not subject to the limitations of filters-based systems. By measuring a definitive spectrum, simultaneously at all points in a sample, SpectraCube TM has the potential to revolutionize FISH by enabling the detection and separation of a large number of spectrally and spatially overlapping fluorescent DNA probes, and by eliminating sample autofluorescence. Details of the Applied Spectral Imaging SD200 spectral bio-imaging system and the results of a six-color FISH measurement will be discussed.