Abstract
Purpose: Our purpose was to evaluate the utility of spectral imaging for multicolor, multichromosome enumeration in human interphase cell nuclei.
Methods: Chromosome-specific probes labeled with different fluorochromes or nonfluorescent haptens were obtained commercially or prepared in-house. Metaphase spreads, interphase lymphocytes, or blastomeres cells were hybridized with either 7 or 11 distinctly different probes. Following 46 hr of hybridization, slides were washed and detected using either a filter-based quantitative image processing system (QUIPS) developed in-house or a commercial spectral imaging system.
Results: The filter-based fluorescence microscope system is preferred for simultaneous detection of up to seven chromosome targets because of its high sensitivity and speed. However, this approach may not be applicable to interphase cells when 11 or more targets need to be discriminated. Interferometer-based spectral imaging with a spectral resolution of approximately 10 nm allows labeling of chromosome-specific DNA probes with fluorochromes having greatly overlapping emission spectra. This leads to increases in the number of fluorochromes or fluorochrome combinations available to score unambiguously chromosomes in interphase nuclei.
Conclusions: Spectral imaging provides a significant improvement over conventional filter-based microscope systems for enumeration of multiple chromosomes in interphase nuclei, although further technical development is necessary in its application to embryonic blastomeres. When applied to preconception/preimplantation genetic diagnosis, presently available probes for spectral imaging are expected to detect abnormalities responsible for 70–80% of spontaneous abortions caused by chromosomal trisomies.
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REFERENCES
Epstein CJ: The Consequences of Chromosome Imbalance: Principles, Mechanisms, and Models. Cambridge, New York, Cambridge University Press, 1986
Hassold TJ, Jacobs PA: Trisomy in man. Annu Rev Genet 1984;18:69-97
Hassold T, Hunt P, Sherman S: Trisomy in humans: Incidence, origin and etiology. Curr Opion Genet Dev 1993;1:398-403
Jacobs PA, Hassold TJ: The origin of numerical chromosome abnormalities. Adv Genet 1995;33:101-133
Munné S, Weier H-UG: Simultaneous enumeration of chromosomes X, Y, 18, 13, and 21 in interphase cells for preimplantationgenetic diagnosis of aneuploidy. Cytogenet Cell Genet 1996;75:264-270
Schröck E, du Manoir S, Veldman T, Schoell B, Wienberg J, Fergueson-Smith M, Ning Y, Ledbetter D, Bar-Am I, Soenksen D, Garini Y, Ried T: Multicolor spectral karyotyping of human chromosomes. Science 1996;273:494-497
Weier H-UG, Wang M, Mullikin JC, Zhu Y, Cheng JF, Greulich KM, Benismon A, Gray JW: Quantitative DNA fiber mapping. Hum Mol Genet 1995;4:1903-1910
Jossart GH, O'Brien B, Cheng JF, Tong Q, Jhiang SM, Duh Q, Clark OH, Weier H-UG: A novel multicolor hybridization scheme applied to localization of a transcribed sequence (D10S170/H4) and deletion mapping in the thyroid cancer cell line TPC-1. Cytogenet Cell Genet 1996;75:254-257
Mascio LN, Verbeek PW, Sudar D, Kuo WL, Gray JW: Semiautomated DNA probe mapping using digital imaging microscopy: I. System development. Cytometry 1995;19:51-69
Harper M, Saunders G: Localization of single copy DNA sequences on G-banded human chromsomes by in situ hybridization. Chromosoma 1981;83:431-439
Munné S, Grifo J, Cohen J, Weier H-UG: Mosaicism and aneuploidy in arrested human preimplantation embryos: A multiple probe fluorescence in situ hybridization (FISH) study. Am J Hum Genet 1994;55:150-159
Moyzis RK, Albright KL, Bartholdi MF, Cram LS, Deaven LL, Hildebrand CE, Joste NE, Longmire JL, Meyne J, Schwarzacher-Robinson T: Human chromosome-specific repetitive DNA sequences: Novel markers for genetic analysis. Chromosoma 1987;95:375-386
Weier H-UG, Miller BM, Yu LC, Fuscoe JC: PCR cloning of a repeated DNA fragment from chinese hamster ovary (CHO) cell X chromosomes and mapping by fluorescence in situ hybridization. DNA Sequence 1993;4:47-51
Cassel MJ, Munné S, Fung J, Weier H-UG: Carrier-specific breakpoint-spanning DNA probes for pre-implantation genetic diagnosis [PGD] in interphase cells. Hum Reprod 1997;12:101-109
Fung J, Munné S, Duell T, Weier H-UG: Rapid cloning of translocation breakpoints: from blood to YAC in 50 days. J Biochem Mol Biol Biophys 1998;1:181-192
Weier H-U, Polikoff D, Fawcett JJ, Greulich KM, Lee K-H, Cram S, Chapman VM Gray JW: Generation of five high complexity painting probe libraries from flow sorted mouse chromosomes. Genomics 1994;24:641-644
Kuo WL, Tenjin H, Segraves R, Pinkel D, Golbus MS, Gray J: Detection of aneuploidy involving chromosomes 13, 18, or 21, by fluorescence in situ hybridization (FISH) to interphase and metaphase amniocytes. Am J Hum Genet 1991;49:112-119
Dauwerse JG, Wiegant J, Raap AK, Breuning AK, van Ommen GJB: Multiple colors by fluorescence in situ hybridization using ratio-labeled DNA probes create a molecular karyotype. Hum Mol Genet 1992;1:593-598
Munné S, Dailey T, Finkelstein M, Weier HUG: Reduction in overlaps and missing signals in interphase-FISH. J Assist Reprod Genet 1996;13:149-156
Spurbeck JL, Zinsmeister AR, Meyer KJ, Jalai SM: Dynamics of chromosome spreading. Am J Med Genet 1996;61:387-393
Munné S, Weier HUG, Grifo JA, Cohen J: Chromosome mosaicism in human embryos. Biol Reprod 1994;51:373-379
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Fung, J., Hyun, W., Dandekar, P. et al. Spectral Imaging in Preconception/Preimplantation Genetic Diagnosis of Aneuploidy: Multicolor, Multichromosome Screening of Single Cells. J Assist Reprod Genet 15, 323–330 (1998). https://doi.org/10.1023/A:1022508930762
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DOI: https://doi.org/10.1023/A:1022508930762