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High resolution multicolor fluorescence in situ hybridization using cyanine and fluorescein dyes: Rapid chromosome identification by directly fluorescently labeled alphoid DNA probes

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Abstract

We tested DNA probes directly labeled by fluorescently labeled nucleotides (Cy3-dCTP, Cy5-dCTP, FluorX-dCTP) for high resolution uni- and multicolor detection of human chromosomes and analysis of centromeric DNA organization by in situ hybridization. Alpha-satellite DNA probes specific to chromosomes 1, 2, 3, 4 + 9, 5 + 19, 6, 7, 8, 10, 11, 13 + 21, 14 + 22, 15, 16, 17, 18, 20, 22, X and Y were suitable for the accurate identification of human chromosomes in metaphase and interphase cells. Cy3-labeled probes had several advantages: (1) a high level of fluorescence (5–10 times more compared with fluorescein-labeled probes); (2) a low level of fluorescence in solution, allowing the detection of target chromosomes in situ during hybridization without the washing of slides; and (3) high resistance to photobleaching during prolonged (1-2 h) exposure to strong light, thus allowing the use of a high energy mercury lamp or a long integration time during image acquisition in digital imaging microscopy for the determination of weak signals. For di- and multicolor fluorescence in situ hybridization (FISH), we successfully used different combinations of directly fluorophorated probes with preservation of images by conventional microscopy or by digital imaging microscopy. FluorX and Cy3 dyes allowed the use of cosmid probes for mapping in a one-step hybridization experiment. Cyanine-labeled fluorophorated DNA probes offer additional possibilities for rapid chromosome detection during a simple 15-min FISH procedure, and can be recommended for basic research and clinical studies, utilizing FISH.

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Authors and Affiliations

  1. National Research Centre of Mental Health, Russian Academy of Medical Sciences, Moscow, Zagorodnoe sh. 2., 113152, Moscow, Russia

    Yuri B. Yurov & Ilia V. Soloviev

  2. Insitute of Pediatrics and Children Surgery, Russian Ministry of Health, Moscow, Russia

    Yuri B. Yurov & Svetlana G. Vorsanova

  3. Institut de Biologie, INSERM U249-CNRS, 9008, Montpellier, France

    Yuri B. Yurov, Svetlana G. Vorsanova, Bertrand Marcais & Gerard Roize

  4. Biological Detection Systems, Inc., Pittsburgh, PA, USA

    Robert Lewis

Authors
  1. Yuri B. Yurov
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  2. Ilia V. Soloviev
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  3. Svetlana G. Vorsanova
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  4. Bertrand Marcais
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  5. Gerard Roize
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  6. Robert Lewis
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Yurov, Y.B., Soloviev, I.V., Vorsanova, S.G. et al. High resolution multicolor fluorescence in situ hybridization using cyanine and fluorescein dyes: Rapid chromosome identification by directly fluorescently labeled alphoid DNA probes. Hum Genet 97, 390–398 (1996). https://doi.org/10.1007/BF02185780

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  • DOI: https://doi.org/10.1007/BF02185780

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