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  • 1
    Electronic Resource
    Electronic Resource
    Somatic pairing of chromosome 1 centromeres in interphase nuclei of human cerebellum (1989)
    Springer
    Human genetics 〈Berlin〉 83 (1989), S. 231-234 
    Details
    ISSN: 1432-1203
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Interphase nuclei isolated from paraffin-embedded tissue of four normal brains were hybridized with biotinated repetitive DNA probes specific for the (peri) centromeric regions of chromosomes 1 and 7. Hybridization results were visualized with a peroxidase-DAB system after which the number of specific signals per nucleus was counted using bright field microscopy. Using the probe specific for chromosome 7 (p7t1), both the cerebral and the cerebellar samples showed 2 spots in 82% and 83%, respectively, of the nuclei. In situ hybridization with the chromosome 1 probe (pUC1. 77) showed two spots in 69% of the cerebral nuclei. In cerebellar samples, hybridization with pUC1.77 resulted in only one large spot per nucleus in 82% of the cells. The average spot size in nuclei with one signal was about 1.6 times as large as that in nuclei with two signals. These observations suggest that the single large spot in the cerebellar cells is not the result of monosomy of chromosome 1 but that it reflects somatic pairing of the two chromosome 1 centromeres. Based on the size and the fraction of nuclei with one large spot, the small granular neuron is the most likely candidate. The difference between cerebral and cerebellar samples indicates that this somatic pairing of chromosome 1 is a cell-type-dependent phenomenon.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00285162
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  • 2
    Electronic Resource
    Electronic Resource
    Anti-DNA.RNA sera (1984)
    Springer
    Histochemistry and cell biology 81 (1984), S. 517-520 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary The specificity of anti-nucleic acid antisera can immunocytochemically be evaluated with test systems which apply various nucleic acids immobilized to inert matrices. When using polyrA.polyrU as a model compoud for dsRNA, it is important to prevent the formation of the triple stranded form polyrA.(polyrU)2. Anti-dsRNA antibodies which, when tested with the correct test system, proved to be present in an earlier described anti-DNA.RNA serum, could be removed by adsorption. By cytofluorometric comparison of the immunofluorescence signals obtained with the anti-dsRNA containing serum and the absorbed serum, it could be shown that the anti-dsRNA antibodies do not contribute to the specific signals measured after in situ hybridization. Repetitive incubations with the anti-DNA.RNA serum led to a considerable increase in immunofluorescence signal.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00489529
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  • 3
    Electronic Resource
    Electronic Resource
    Localization properties of fluorescence cytochemical enzyme procedures (1986)
    Springer
    Histochemistry and cell biology 84 (1986), S. 317-321 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Fluorescence enzyme cytochemical procedures will contribute significantly to biomedical problems where knowledge of the enzymic composition of individual cells is important. Compared with the number of absorbance enzyme cytochemical methods, relatively few fluorescence procedures have been reported. In this paper, the merits of the described methods are discussed. A distinction is made between methods with and without a capture reaction. Only a few methods satisfy the requirement of accurate localization of the final product and high signal to noise ratios. Thus, there still is a need for valid fluorescence cytochemical enzyme methods. It is concluded that the bottle neck for valid fluorescence cytochemical enzyme methods is the development of efficient fluorogenic capture reactions for the primary enzyme products.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00482956
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  • 4
    Electronic Resource
    Electronic Resource
    Bi-color detection of two target DNAs by non-radioactive in situ hybridization (1986)
    Springer
    Histochemistry and cell biology 85 (1986), S. 1-4 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary A non-radioactive in situ hybridization technique is described which allows the simultaneous detection of different DNA sequences. To demonstrate the feasibility of the proccdure, metaphases and interphase nuclei of a human-mouse somatic cell hybrid were simultaneously hybridized with mercurated total human DNA and a biotinylated mouse satellite DNA probe. After the hybridization, the probes were detected immunocytochemically using two different and independent affinity systems. By this approach we visualized the two DNA target sequences in metaphase chromosomes and in interphase nuclei with FITC and TRITC fluorescence, or blue (alkaline phosphatase) and brown (peroxidase) precipitated enzyme products. This method not only allows detection of intact chromosomes but also the visualization of rearrangements between parts of human and mouse chromosomes. Furthermore, the technique demonstrates the high topological resolution of nonradioactive in situ hybridizations.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00508646
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  • 5
    Electronic Resource
    Electronic Resource
    In situ hybridization as a tool to study numerical chromosome aberrations in solid bladder tumors (1988)
    Springer
    Histochemistry and cell biology 89 (1988), S. 307-316 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary Methods for single- and double-target in situ hybridization (ISH) to, cells isolated from solid transitional cell carcinomas (TCC's) of the urinary bladder are described. Single cell suspensions were prepared from solid tumors of the urinary bladder by mechanical disaggregation and fixed in 70% ethanol. Using two DNA probes specific for the centromeres of chromosomes #1 and #18, ISH procedures were optimized for these samples. Human lymphocytes and cells from the T24 bladder tumor cell line were used as controls. In lymphocyte nuclei and metaphase chromosome spreads, ISH showed two major spots for each of the probes. About 80% of the nuclei from T24 cells showed three spots for both the chromosome #1 and #18 specific probes. When nuclei from TCC's were analyzed, often the number of spots for chromosome #1, and to a lesser extent for chromosome #18, differed from the number expected on basis of flow cytometric ploidy measurements. The double target-ISH method in all cases allowed the correlation of numerical aberrations for chromosomes #1 and #18 in one and the same cell. By such analyses a profound heterogeneity in chromosome number was detected in most tumors. In order to optimize the reproductbility of the method and the interpretation of the ISH-signals, criteria for their analysis have been determined. This procedure can now be applied on a routine basis to solid tumor specimens.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00500631
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  • 6
    Electronic Resource
    Electronic Resource
    Early replication signals in nuclei of Chinese hamster ovary cells (1990)
    Springer
    Histochemistry and cell biology 94 (1990), S. 435-440 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Summary DNA replication sites generally known as replicon domains were resolved as individual replication signals in interphase nuclei of permeabilized Chinese hamster ovary cells by immunofluorescent microscopy. Biotin-11-dUTP was utilized as a tool to label newly replicated DNA in permeable cells and to study the distribution of nascent DNA in pulselabel and in pulsechase experiments. Active sites of DNA replication were visualized in exponentially growing cells and in synchronized cultures throughout the S phase. Fluorescent images of replication sites were analyzed by standard fluorescense microscopy and in three dimensions by confocal laser scanning microscopy. The rapid increase in number of discrete foci of newly replicated DNA is an indication that DNA synthesis starts at limited number of sites in mammalian nuclei rather than at thousands of foci at the same time.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00266452
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  • 7
    Electronic Resource
    Electronic Resource
    Oestradiol, a new hapten for detecting nucleic acid sequences by FISH (1997)
    Springer
    Histochemistry and cell biology 108 (1997), S. 359-364 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  Oestradiol has been conjugated to allylamine-dUTP with an 11-atom spacer to allow enzymatic incorporation of the label into DNA sequences. In a comparative DNA and mRNA FISH study we have used DNA probes that were either labelled with digoxigenin, biotin or oestradiol. Results show that oestradiol-labelled probes can detect DNA and RNA sequences in FISH equally well as digoxigenin- and biotin-labelled probes. Further, no crossreactivity between the various hapten-specific antibodies and the three haptens were observed. Binding of the rabbit anti-oestradiol antibody to endogenous oestrogen in various tissues was not observed under the conditions tested. In view of the increasing demands for multi-colour DNA and mRNA FISH applications, oestradiol is a welcome addition to the collection of haptens employed in FISH.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004180050176
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  • 8
    Electronic Resource
    Electronic Resource
    EDITORIAL (1997)
    Springer
    Histochemistry and cell biology 108 (1997), S. 289-289 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004180050167
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  • 9
    Electronic Resource
    Electronic Resource
    Fluorescence in situ hybridization using horseradish peroxidase-labeled oligodeoxynucleotides and tyramide signal amplification for sensitive DNA and mRNA detection (1998)
    Springer
    Histochemistry and cell biology 110 (1998), S. 431-437 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  We have used horseradish peroxidase-labeled 40 mer oligodeoxynucleotides (HRP-ODNs) specific for the human cytomegalovirus immediate early gene (HCMV-IE) and a novel dinitrophenol-tyramide signal amplification reagent (DNP-TSA plus) to evaluate their utility in fluorescence in situ hybridization (FISH). For DNA FISH, single or cocktails of HRP-ODNs were hybridized to metaphase chromosomes of rat 9G cells which, as determined by DNA fiber FISH, carry an integrated tandem repeat of 50–60 copies of the HCMV-IE gene. With one layer of DNP-TSA plus deposition and subsequent detection with a fluorochrome-conjugated antibody, four HRP-ODNs were needed to detect the HCMV-IE integration site. When employing two TSA amplification rounds, one HRP-ODN was sufficient for obtaining a strong signal of the integrated gene cluster, indicating that 50–60 HRP molecules can be detected with ease. In addition to DNA FISH, we report here the first use of HRP-ODN probes for mRNA detection by FISH. A single HRP-ODN and one DNP-TSA plus step resulted in clear visualization of the HCMV-IE gene transcripts in rat 9G cells induced for HCMV-IE expression by cycloheximide. Two TSA detection steps enhanced signal intensities even further. Parallel experiments with hapten-labeled ODN and cDNA probes and conventional detection methods illustrated the superiority of the HRP-ODN/TSA approach in DNA and RNA FISH.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004180050304
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  • 10
    Electronic Resource
    Electronic Resource
    Synthesis and purification of horseradish peroxidase-labeled oligonucleotides for tyramide-based fluorescence in situ hybridization (2000)
    Springer
    Histochemistry and cell biology 113 (2000), S. 175-180 
    Details
    ISSN: 1432-119X
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract  A method is presented to conjugate horseradish peroxidase (HRP) to oligodeoxynucleotides for fluorescence in situ hybridization assays employing tyramide signal amplification (TSA). HRP is covalently bound to the oligonucleotide by thiol ether linkage and purified by high-performance liquid chromatography. With TSA detection, a single HRP-labeled oligonucleotide probe is sufficient for in situ detection of clustered DNA repeat sequences with a degree of repetition between 20 and 50.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/s004180050436
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