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Journal ArticleDOI

Multicolor Spectral Karyotyping of Human Chromosomes

26 Jul 1996-Science (American Association for the Advancement of Science)-Vol. 273, Iss: 5274, pp 494-497
TL;DR: Whole-genome scanning by spectral karyotyping allowed instantaneous visualization of defined emission spectra for each human chromosome after fluorescence in situ hybridization, and all human chromosomes were simultaneously identified.
Abstract: The simultaneous and unequivocal discernment of all human chromosomes in different colors would be of significant clinical and biologic importance. Whole-genome scanning by spectral karyotyping allowed instantaneous visualization of defined emission spectra for each human chromosome after fluorescence in situ hybridization. By means of computer separation (classification) of spectra, spectrally overlapping chromosome-specific DNA probes could be resolved, and all human chromosomes were simultaneously identified.
Citations
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Journal ArticleDOI
25 Sep 1998-Science
TL;DR: Semiconductor nanocrystals prepared for use as fluorescent probes in biological staining and diagnostics have a narrow, tunable, symmetric emission spectrum and are photochemically stable.
Abstract: Semiconductor nanocrystals were prepared for use as fluorescent probes in biological staining and diagnostics. Compared with conventional fluorophores, the nanocrystals have a narrow, tunable, symmetric emission spectrum and are photochemically stable. The advantages of the broad, continuous excitation spectrum were demonstrated in a dual-emission, single-excitation labeling experiment on mouse fibroblasts. These nanocrystal probes are thus complementary and in some cases may be superior to existing fluorophores.

8,542 citations

Journal ArticleDOI
TL;DR: This review looks at current methods for preparing QD bioconjugates as well as presenting an overview of applications, and concludes that the potential of QDs in biology has just begun to be realized and new avenues will arise as the ability to manipulate these materials improves.
Abstract: One of the fastest moving and most exciting interfaces of nanotechnology is the use of quantum dots (QDs) in biology. The unique optical properties of QDs make them appealing as in vivo and in vitro fluorophores in a variety of biological investigations, in which traditional fluorescent labels based on organic molecules fall short of providing long-term stability and simultaneous detection of multiple signals. The ability to make QDs water soluble and target them to specific biomolecules has led to promising applications in cellular labelling, deep-tissue imaging, assay labelling and as efficient fluorescence resonance energy transfer donors. Despite recent progress, much work still needs to be done to achieve reproducible and robust surface functionalization and develop flexible bioconjugation techniques. In this review, we look at current methods for preparing QD bioconjugates as well as presenting an overview of applications. The potential of QDs in biology has just begun to be realized and new avenues will arise as our ability to manipulate these materials improves.

5,875 citations


Cites background from "Multicolor Spectral Karyotyping of ..."

  • ...1a ) and low photobleaching thresholds, have limited their effectiveness in long-term imaging and 'multiplexing' (simultaneous detection of multiple signals) without complex instrumentation and processin...

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Journal Article
TL;DR: The identification and purification of a cancer stem cell from human brain tumors of different phenotypes that possesses a marked capacity for proliferation, self-renewal, and differentiation is reported.
Abstract: Most current research on human brain tumors is focused on the molecular and cellular analysis of the bulk tumor mass. However, there is overwhelming evidence in some malignancies that the tumor clone is heterogeneous with respect to proliferation and differentiation. In human leukemia, the tumor clone is organized as a hierarchy that originates from rare leukemic stem cells that possess extensive proliferative and self-renewal potential, and are responsible for maintaining the tumor clone. We report here the identification and purification of a cancer stem cell from human brain tumors of different phenotypes that possesses a marked capacity for proliferation, self-renewal, and differentiation. The increased self-renewal capacity of the brain tumor stem cell (BTSC) was highest from the most aggressive clinical samples of medulloblastoma compared with low-grade gliomas. The BTSC was exclusively isolated with the cell fraction expressing the neural stem cell surface marker CD133. These CD133+ cells could differentiate in culture into tumor cells that phenotypically resembled the tumor from the patient. The identification of a BTSC provides a powerful tool to investigate the tumorigenic process in the central nervous system and to develop therapies targeted to the BTSC.

4,899 citations


Cites methods from "Multicolor Spectral Karyotyping of ..."

  • ...Spectral images were acquired and analyzed with an SD 200 Spectral Bio-imaging System (ASI Ltd., MigdalHaemek, Israel) attached to a Zeiss Axioplan 2 microscope (Carl Zeiss, Toronto, Ontario, Canada), and analyzed using SKYVIEW (ver....

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  • ...Detailed SKY analysis was possible in 8 metaphases, and all of the cells had an identical clonally abnormal karyotype....

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  • ...SKY was performed on tumor metaphase cells according to the manufacturer’s instructions (ASI, Carlsbad, CA) and as published previously (8)....

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  • ...3 The abbreviations used are: BTSC, brain tumor stem cell; TSM, tumor sphere medium; EGF, epidermal growth factor; bFGF, basic fibroblast growth factor; FBS, fetal bovine serum; PDGFR, platelet-derived growth factor receptor; SKY, spectral karyotyping; SFM, serum-free medium; GFAP, glial fibrillary acidic protein....

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  • ...We also performed cytogenetic analysis and SKY (8) using metaphase preparations obtained directly from cultured tumor spheres from a medulloblastoma (Patient 7; Fig....

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Journal ArticleDOI
TL;DR: Applications in Theranostics Guanying Chen,*,†,‡ Hailong Qiu,*,‡ and Xiaoyuan Chen.
Abstract: Applications in Theranostics Guanying Chen,*,†,‡ Hailong Qiu,†,‡ Paras N. Prasad,*,‡,§ and Xiaoyuan Chen* †School of Chemical Engineering and Technology, Harbin Institute of Technology, Harbin, Heilongjiang 150001, China ‡Department of Chemistry and the Institute for Lasers, Photonics, and Biophotonics, University at Buffalo, State University of New York, Buffalo, New York 14260, United States Department of Chemistry, Korea University, Seoul 136-701, Korea Laboratory of Molecular Imaging and Nanomedicine, National Institute of Biomedical Imaging and Bioengineering, National Institutes of Health, Bethesda, Maryland 20892-2281, United States

1,994 citations

Journal ArticleDOI
31 Oct 1996-Nature
TL;DR: In this article, it was shown that light emission from single fluorescing nanocrystals of cadmium selenide under continuous excitation turns on and off intermittently with a characteristic timescale of about 0.5 seconds.
Abstract: SEMICONDUCTOR nanocrystals offer the opportunity to study the evolution of bulk materials properties as the size of a system increases from the molecular scale1,2. In addition, their strongly size-dependent optical properties render them attractive candidates as tunable light absorbers and emitters in optoelectronic devices such as light-emitting diodes3,4 and quantum-dot lasers5,6, and as optical probes of biological systems7. Here we show that light emission from single fluorescing nanocrystals of cadmium selenide under continuous excitation turns on and off intermittently with a characteristic timescale of about 0.5 seconds. This intermittency is not apparent from ensemble measurements on many nanocrystals. The dependence on excitation intensity and the change in on/off times when a passivating, high-bandgap shell of zinc sulphide encapsulates the nanocrystal8,9 suggests that the abrupt turning off of luminescence is caused by photo-ionization of the nanocrystal. Thus spectroscopic measurements on single nanocrystals can reveal hitherto unknown aspects of their photophysics.

1,757 citations

References
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Journal ArticleDOI
TL;DR: The data suggest that multiplex-fluorescence in situ hybridization (M-FISH) could have wide clinical utility and complement standard cytogenetics, particularly for the characterization of complex karyotypes.
Abstract: We have developed epifluorescence filter sets and computer software for the detection and discrimination of 27 different DNA probes hybridized simultaneously. For karyotype analysis, a pool of human chromosome painting probes, each labelled with a different fluor combination, was hybridized to metaphase chromosomes prepared from normal cells, clinical specimens, and neoplastic cell lines. Both simple and complex chromosomal rearrangements could be detected rapidly and unequivocally; many of the more complex chromosomal abnormalities could not be delineated by conventional cytogenetic banding techniques. Our data suggest that multiplex-fluorescence in situ hybridization (M-FISH) could have wide clinical utility and complement standard cytogenetics, particularly for the characterization of complex karyotypes.

1,333 citations

Journal ArticleDOI
TL;DR: The air-drying method gives improved spreading of the chromosomes and less cell breakage compared with squash techniques, suitable for stages of male meiosis in which the chromosomes are condensed.
Abstract: A suspension is made in isotonic (2.2%) sodium citrate solution from the contents of the tubules from a whole testis or a testicular biopsy specimen. The germinal cells are sedimented by centrifuging, leaving most of the sperm in the supernatant fluid, which is discarded. The cells are resuspended in hypo-tonic (1%) sodium citrate solution and left to stand at room temperature for 12 minutes, after which they are sedimented again and fixed as a concentrated suspension in a mixture of 3 parts absolute ethyl alcohol to 1 part glacial acetic acid plus a trace of chloroform. Two quick changes into fresh fixative follow. Air-dried preparations are made from the final fixed suspension and stained in lactic-acetic-orcein. The method is suitable for stages of male meiosis in which the chromosomes are condensed. Its principle advantage is the separation of the clumps of spermatogonia and spermatocytes into individual cells which are randomly dispersed over the preparations. Compared with squash techniques, the air-drying method gives improved spreading of the chromosomes and less cell breakage.

1,032 citations

Journal ArticleDOI
TL;DR: Comparative genomic hybridization was applied to 5 breast cancer cell lines and 33 primary tumors to discover and map regions of the genome with increased DNA-sequence copy-number, indicating that these chromosomal regions may contain previously unknown genes whose increased expression contributes to breast cancer progression.
Abstract: Comparative genomic hybridization was applied to 5 breast cancer cell lines and 33 primary tumors to discover and map regions of the genome with increased DNA-sequence copy-number. Two-thirds of primary tumors and almost all cell lines showed increased DNA-sequence copy-number affecting a total of 26 chromosomal subregions. Most of these loci were distinct from those of currently known amplified genes in breast cancer, with sequences originating from 17q22-q24 and 20q13 showing the highest frequency of amplification. The results indicate that these chromosomal regions may contain previously unknown genes whose increased expression contributes to breast cancer progression. Chromosomal regions with increased copy-number often spanned tens of Mb, suggesting involvement of more than one gene in each region.

772 citations

Journal ArticleDOI
TL;DR: This study shows that flow sorting of aberrant chromosomes and chromosome painting can be used as a rapid aid to cytogenetic analysis, particularly in cases of difficult karyotypes, such as tumours.
Abstract: A novel polymerase chain reaction (PCR) technique has been combined with chromosome flow sorting to characterise two lymphoblastoid cell lines and one medullary thyroid carcinoma cell line carrying translocations close to the locus for multiple endocrine neoplasia type 2A (MEN 2A). Five hundred copies of the derivative chromosome(s) were flow sorted from each cell line and amplified by degenerate oligonucleotide-primed-polymerase chain reaction (DOP-PCR). This generated pools of DNA sequences corresponding to the abnormal chromosomes, which were then used as probes in fluorescence in situ hybridisation (FISH) experiments on normal metaphase cells. The resultant chromosome paints revealed the portions of the normal chromosomes related to those involved in the translocations. By this technique, translocation breakpoints in bands p15, q11.2, and q21 of chromosome 10 were defined in the above cell lines, in two cases refining previous cytogenetic data. This study shows that flow sorting of aberrant chromosomes and chromosome painting can be used as a rapid aid to cytogenetic analysis, particularly in cases of difficult karyotypes, such as tumours. Furthermore, the DOP-PCR technique described here will have applications to other areas of genome analysis, such as cloning of new markers; its design will allow a general and representative amplification to occur from any starting DNA in any species.

584 citations

Journal ArticleDOI
TL;DR: The construction of the first complete genetic linkage map of the laboratory rat is reported, identifying 432 markers that show polymorphisms between the SHR and BN rat strains and mapped them in a single SHR × BN F2 intercross.
Abstract: We report the construction of the first complete genetic linkage map of the laboratory rat. By testing 1171 simple sequence length polymorphisms (SSLPs), we have identified 432 markers that show polymorphisms between the SHR and BN rat strains and mapped them in a single (SHR × BN) F2 intercross. The loci define 21 large linkage groups corresponding to the 21 rat chromosomes, together with a pair of nearby markers on chromosome 9 that are not linked to the rest of the map. Because 99.5% of the markers fall into one of the 21 large linkage groups, the maps appear to cover the vast majority of the rat genome. The availability of the map should facilitate whole genome scans for genes underlying qualitative and quantitative traits relevant to mammalian physiology and pathobiology.

478 citations