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Results for “In Situ Hybridization, Fluorescence”

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At least 19 recordsLinked to original sources
PubMed2001

Complete karyotype characterization of the K562 cell line by combined application of G-banding, multiplex-fluorescence in situ hybridization, fluorescence in situ hybridization, and comparative genomic hybridization.

This study combines conventional cytogenetics, fluorescence in situ hybridization (FISH), multiplex-FISH and comparative genomic hybridization (CGH). In applying this multimodal approach on the human leukemia cell line K562, the chromosome composition was refined in detail and compared with data from the literature. A hypotriploid karyotype with a modal chromosome number of 67, and 21 unique marker chromosomes were identified. The classification of six markers was identical to published data and the composition of five further markers from the literature could be fully clarified for the first time. The composition of another five markers, which have been interpreted in divergent ways in different studies, were elucidated without doubt. Finally, five new markers of our study seem to have no equivalents in former studies, very likely due to limitations of conventional cytogenetics. The combinatory application of complementary techniques as shown in this study will be very useful to provide the basis of a refined genotype analysis on the chromosomal level.

Chromosome Mapping↗
PubMed2001

[Localization and position effect of human APPSWE gene on transgenic mice by means of in situ fluorescence hybridization].

In situ fluorescence hybridization (FISH) was used to study the integration and localization of human APPSWE gene on chromosomes of transgenic mice. It was found that in 2 founder mice, 34 and 36 metaphases of 80 metaphases observed had shown obvious specific hybridizing signals, the detection rates were 42.5% and 45%, respectively; meanwhile, in 1 F1 and 1 F2 mice, 33 and 30 metaphases out of 100 metaphases observed had specific hybridizing signals, the detection rates were 33% and 30%, respectively. The transgenes were localized on chromosome 8, 1, 17 and 2. The results indicated that transgene APPSWE were integrated stably into the chromosomes of transgenic mice and could be transmitted to offsprings through germ cells. The transgenes were randomly integrated into multiple sites of mice chromosomes. Meanwhile, the phenotypes of transgenic mice were also studied and different integrated sites had obvious effects on the pheonotypes.

Amyloid beta-Protein Precursor↗
PubMed1997

Characterization of a de novo unbalanced chromosome rearrangement by comparative genomic hybridization and fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) has proven useful for the identification of chromosomal material of unknown origin. More recently, comparative genomic hybridization (CGH) has been used to identify deletions and amplifications, particularly in neoplastic samples. Here, we describe the combined use of CGH and FISH to identify the origin of a de novo unbalanced translocation in a newborn with multiple congenital anomalies. GTG banding of metaphases from cultured lymphocytes showed an unbalanced karyotype, with extra material on a chromosome 5: 46,XX,add(5)(q35). Parental karyotypes were both normal. CGH revealed the additional material was from distal 11q (11q23-->'qter). This finding was confirmed by FISH with a whole chromosome paint for chromosome 11. Based on the CGH and FISH analyses, the proband's karyotype was therefore 46,XX,der(5)t(5;11)(q35.2; q23.2).ish der(5)(wcp11+). This case demonstrates the efficient use of CGH and confirmatory FISH for the identification of chromosomal material of unknown origin.

Abnormalities, Multiple↗
PubMed2006

New approaches to fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) is a nonisotopic labeling and detection method that provides a direct way to determine the relative location or copy number of specific DNA sequences in nuclei or chromosomes. With recent advancements, this technique has found increased application in a number of research areas, including cytogenetics, prenatal diagnosis, cancer research and diagnosis, nuclear organization, gene loss and/or amplification, and gene mapping. The availability of different types of probe and the increasing number of FISH techniques has made it a widespread and diversely applied technology. Multicolor karyotyping by multicolor FISH and spectral karyotyping interphase FISH and comparative genomic hybridization allow genetic analysis of previously intractable targets. We present a brief overview of FISH technology and describe in detail methods of probe labeling and detection for different types of tissue sample, including microdissected nuclei from formalin-fixed paraffin-embedded tissue sections.

Cell Cycle↗
PubMed2001

A case of adult T-cell leukaemia/lymphoma characterized by multiplex-fluorescence in situ hybridization, comparative genomic hybridization, fluorescence in situ hybridization and cytogenetics.

Adult T-cell leukaemia/lymphoma (ATLL) is a neoplasm of mature helper (CD4) T lymphocytes. Little is known, however, about the chromosome aberrations associated with the pathogenesis of this malignancy. Using molecular cytogenetic techniques we, therefore, investigated a 44-year-old man who had a 7-year history of ATLL with cutaneous involvement mimicking primary cutaneous T-cell lymphoma. Conventional cytogenetics revealed gross chromosomal changes with chromosome numbers ranging from 71 to 82. There were structural abnormalities of chromosomes 7 and 9, partial deletions of chromosomes 1, 3, 5 and 6, and loss of chromosomes 2, 4, 9, 11--14, 21 and 22. Multiplex-fluorescence in situ hybridization (M-FISH) identified two derivative chromosomes, der(6)t(6;7)(q16;q21) and der(7)t(6;7)(q16;q21)ins(6;12)(q2?;?), and a deletion of chromosome 1p. Conventional FISH confirmed the M-FISH findings. Comparative genomic hybridization of the blood revealed gains of DNA copy number at 1q12--25, 6p24--25, 9p23, 16p13--q13, 17q11--21, 19p13 and 20q13 and loss at 11p15 while lymph nodes showed gains at 3p22--24, 3q27--29, 7q36 and 15q26 and losses at 2p24--25, 2q37, 10p14--15, 11p15, 13q33--34 and 16p13.3. No DNA copy number changes were seen in a skin lesion. These results show the extent of genetic abnormalities within this malignancy.

Adult↗
PubMed2005

In situ functional gene analysis: recognition of individual genes by fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) using specific probes certainly is one of the most commonly applied molecular techniques in microbial ecology. Monitoring of community composition and dynamics can be combined with localization and identification of individual cells in situ. However, the resolution power of the method is limited by the need for high target numbers per cell. Apart from standard targets (ribosomal RNAs), mRNAs could be used successfully for in situ visualization in some cases. A new promising variant of in situ hybridization could be established that should provide access to any low copy number nucleic acid targets, such as chromosomal genes. The recognition of individual genes by FISH (RING-FISH) technology is based on target visualization mediated by polynucleotide probe network formation. The specificity of the approach is provided by intracellular probe-target hybridization. This initial hybridization apparently acts as a focal point for inter-probe hybridization within and mainly in the periphery of the cells. Probe-conferred fluorescence typically appears halo-like around the cells. RING-FISH can be used in combination with conventional oligonucleotide FISH. Thus, genetic potential can be assigned to in situ identified cells.

Escherichia coli↗
PubMed1994

Systemic chimerism in sex-mismatched liver transplant recipients detected by fluorescence in situ hybridization.

Fluorescent in situ hybridization (FISH) is a reliable, rapid, sensitive, and quantitative method for detection of residual host cells following sex-mismatched bone marrow transplantation. Recently, donor-derived long-term multilineage hematopoiesis was detected in a sex-mismatched liver transplant recipient. We therefore assessed chimeric status in 12 patients (F 9, M3), mean age 42.5 years (range 16-57), for a median period of 18 months (range 7-32) following sex-mismatched liver transplantation. Peripheral blood hematolymphoid cells were hybridized with Y- or X-chromosome fluorescently labeled specific probes, and the donor-typed hematopoietic cells were enumerated. In two F recipients 4-5% male hematolymphoid cells were detected in the peripheral blood at 15 and 22 months after sex-mismatched liver transplantation, respectively. These two patients with systemic chimerism suffered from primary biliary cirrhosis and fulminant Wilson's disease before transplantation. One of them had evidence of graft rejection only once during the posttransplant course and the other had no episode of graft rejection. Two other female patients who were found to have approximately 2% male hematolymphoid cells, which is considered to be in the false-positive range, also had no signs of graft rejection during the posttransplant follow-up period. Among the remaining eight patients, in whom systemic chimerism was undetectable, there was at least one episode of acute cellular rejection during the posttransplant period. In summary, the FISH technique enables us to detect systemic chimerism following sex-mismatched liver allografts. Inasmuch as balanced systemic chimerism after organ transplantation is of major importance for self tolerance, our findings may enable us to treat patients after liver transplantation without the need for immunosuppression.

Adult↗
PubMed1996

The use of archival frozen tumor tissue imprint specimens for fluorescence in situ hybridization.

Fluorescence in situ hybridization for the detection of gene amplification or deletion has great promise as a method of providing diagnostic or prognostic information from tumor specimens. Fine needle aspiration samples or tumor tissue imprints are much easier to use and provide better results than paraffin sections, however, these materials are rarely archived and it may take many months to accumulate enough specimens for a study. Archival breast carcinoma tumor tissue samples, some stored frozen for several years, were used to prepare tumor tissue touch imprints on glass microscope slides. The imprints were subjected to fluorescence in situ hybridization analyses utilizing a biotin-dUTP-labeled genomic DNA probe for the cyclin D1 gene (CCND1/PRAD1) and a digoxygenin-labeled chromosome 11 alpha-satellite probe to control for chromosomal copy number. Amplification of CCND1 was easily detectable in frozen tissue imprints. The results indicate that both cytologic morphology and hybridization capacity are well preserved in archived frozen tissue and easily permit its use for in situ hybridization experiments. The ability to use stored frozen tumor tissue for molecular morphologic analysis should allow more rapid assessment of fluorescence in situ hybridization as a potential adjunct for tumor analysis by the surgical pathologist.

Biopsy↗
PubMed1996

Aberrations of chromosomes 9 and 17 in bilharzial bladder cancer as detected by fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH), using repetitive alpha satellite probes to chromosomes 9 and 17, was performed on 27 paraffinembedded bladder specimens from 18 Egyptian patients affected with bilharzial carcinoma. Fluorescence in situ hybridization results in the carcinomas and benign mucosa of patients with schistosomiasis were compared with flow cytometric DNA ploidy and cell cycle assays. These results demonstrate the feasibility of this technique for studies of these archival specimens, and suggest early onset of chromosomal aberrations in histologically benign mucosa of schistosomal cystitis with progression during the development of bladder cancer. On the basis of these data, monosomy 9 may be an early chromosomal change in urothelium of the bilharzia-infested bladder, and a predictor of incipient carcinoma in patients with bilharzial cystitis.

Carcinoma↗
PubMed1996

Detection of numerical chromosome anomalies in interphase cells of ovarian carcinomas using fluorescence in situ hybridization.

Fluorescence in situ hybridization was used in interphase cells of 30 ovarian carcinomas to detect numerical changes in copy number of 13 different centromeres (1, 2, 3, 4, 6, 7, 8, 10, 11, 12, 17, 18, and X). Thirty-seven percent of samples (11/30) were near diploid and demonstrated only minor changes in centromere copy number, involving gain and/or loss of one or a few centromeres. The most common changes included loss of centromeres 4,6, 17, and 18 and gain of centromere 1. The remaining 63% of samples were hyperdiploid and demonstrated a general increase in copy number of most or all centromeres examined. Among these samples, the centromere of chromosome I was most often found to be at higher copy number. Centromeres that were less often at increased copy or deleted within the hyperdiploid samples include centromeres 4, 17, 18, and X. These results suggest that tumor-suppressor genes that are located on chromosomes 4, 6, 17, and 18 may be involved in the development and progression of ovarian cancer.

Adult↗
PubMed1993

Identification of a cryptic t(5;7) reciprocal translocation by fluorescent in situ hybridization.

Fluorescent in situ hybridization (FISH) identified a cryptic balanced reciprocal translocation in the mother of an infant with the cri-duchat syndrome. A biotinylated probe from a flow-sorted chromosome 5 cosmid library was used to show the distal deletion of 5p15.2 in the propositus and a translocation of this segment to the distal end of 7 at 7p21 in his mother. In a subsequent pregnancy, the fetus was shown to have normal chromosomes using the same 5 cosmid library probe and a locus-specific probe derived from the 5p15.3 region. The importance of incorporating FISH into the routine diagnostic laboratory is discussed.

Adult↗
PubMed1994

Improved technique for analysis of formalin-fixed, paraffin-embedded tumors by fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) and specific DNA probes for peri-centromeric repeat regions and unique sequence loci have made it possible to study chromosomal aberrations from interphase tumor nuclei. Large-scale retrospective studies on the prognostic value of interphase cytogenetics would become feasible if these techniques were readily applicable to nuclei from archival formalin-fixed tumor tissues. We describe here an improved technique for interphase FISH analysis of tumors that have been extensively fixed in formalin. The protocol aims at improving probe penetration and hybridization efficiency by inducing chromatin decondensation and swelling of the nuclei with a heat treatment in a 90 degrees C glycerol solution prior to hybridization. Using this cell pretreatment, FISH results on the detection of chromosome copy number aberrations and amplification of the c-erbB-2 oncogene from formalin-fixed, paraffin-embedded tissues were highly concordant with those from fresh tissues. In contrast to previously described methods, separate adjustments of denaturation or proteinase K digestion are not required for each sample. This method facilitates retrospective analyses of large series of tumors and is also useful for applying FISH to routine diagnostic purposes using formalin-fixed material.

Breast Neoplasms↗
PubMed1993

Analysis of restriction enzyme-induced chromosomal aberrations by fluorescence in situ hybridization.

Fluorescence in situ hybridization and Giemsa staining of metaphase chromosomes were used to determine the relative frequencies of symmetric exchange aberrations (translocations) and asymmetric exchange aberrations (rings, dicentrics, and polycentrics) after exposure of human lymphoblastoid cells to restriction enzymes or X-rays. The yield of symmetric exchanges was determined with the use of chromosome-specific probes for human chromosomes 2 or 4, which were hybridized to metaphase chromosomes from cells exposed to the enzymes PvuII, SacI, or XbaI or 3 or 5 Gy of X-rays. The yield of asymmetric exchanges was determined in Giemsa-stained metaphase chromosomes from the same enzyme-treated or irradiated cell population. About 1.5- to 3-fold more symmetric than asymmetric exchanges were induced after restriction enzyme treatment. However, after X-ray treatment the yield of dicentrics relative to the yield of reciprocal translocations was close to the expected 1:1 ratio.

Cell Line↗
PubMed1993

Do terminal deletions of 11q23 exist? Identification of undetected translocations with fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) was performed on bone marrow or peripheral blood cells thought to contain a del(11)(q23q25) from four patients who had acute leukemia or myelodysplasia. Cells from all patients were shown to contain translocations that involved chromosome 6 in three of them. Our data suggest that a large proportion of presumptive del(11)(q23) or del(11)(q23q25) chromosomes may represent previously unidentified translocations that can be detected by FISH.

Acute Disease↗
PubMedDate not supplied

Identification of chromosomal rearrangements in the human myeloid leukemia cell line GF-D8 by dual-colour fluorescence in situ hybridization.

Fluorescence In Situ Hybridization (FISH) studies with chromosome-specific libraries and repetitive probes were performed on the human acute myeloid leukemia cell line GF-D8 in order to define the complex chromosomal rearrangements observed by conventional cytogenetic analysis. Two-colour FISH with whole chromosome painting probes 8 and 11 showed that the add(8) chromosome had an 11-derived region inserted at q24, whereas the add(11) chromosome had an 8-derived region translocated onto q23. It also demonstrated that no normal chromosome 11 is present in GF-D8 cells, since a translocation involving chromosomes 11 and 17q was detected in addition to the add(11). The der(7) chromosome with extra material in its long arm, identified by QFQ and GTG banding, turned out to have a chromosome 15-derived segment translocated to q22. The deletion of 7q was proved to be interstitial, as the 7q-specific telomere as well as a tiny 7-specific band were observed on an unknown chromosome. Fine mapping of the breakpoints involved in the multiple chromosomal rearrangements of the GF-D8 cell line might provide insights into the mechanisms of myeloid leukaemogenesis.

Chromosome Aberrations↗
PubMed1992

Detection of chromosome 17- and X-bearing human spermatozoa using fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) with DNA probes specific to chromosomes 17 and the X has been applied to human ejaculated sperm. After sperm nuclei were decondensed with EDTA and DTT, biotinylated alpha satellite DNA probes TR17 and TRX were separately used on preparations from thirteen healthy donors. After hybridization 96% of sperm were labelled with the TR17 probe and 48% of sperm were labelled with the TRX probe. Frequencies of 0.33% disomic 17 and 0.29% disomic X sperm were found. The frequencies of diploid sperm were assessed as 0.37% using the TR17 probe and 0.20% using the TRX probe which labelled only one half of the sperm; after correcting the result from the X-probe to 0.40% the two frequencies are very similar.

Aneuploidy↗
PubMed1994

Numerical chromosome abnormalities in spermatozoa of fertile and infertile men detected by fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) with single-color chromosome-specific probes was used to study the rates of disomy for chromosome 1, 16, X, and Y in sperm of fertile and infertile subjects. Diploidy rates were studied using a two-color cocktail of probes for chromosomes 17 and 18 in the same sperm samples. Two-color methodology was not available at the outset of the study. A total of 450,580 spermatozoa were studied from 21 subjects (9 fertile, 12 infertile). Significant differences were observed in the disomy rates between chromosomes with the highest frequency observed for chromosome 16 (0.17%) and the lowest for the Y chromosome (0.10%). No differences were observed between fertile and infertile subjects for either diploidy or disomy. Total disomy rates for chromosomes 1, 16, X and Y ranged from 0.34% to 0.84% among infertile subjects, and 0.32% to 0.61% among fertile subjects. Our data suggest that generalized aneuploidy in sperm is not a major contributor to unexplained infertility.

Aneuploidy↗
PubMed1995

Confirmation of the copy number of chromosome 1 in interphase nuclei from paraffin sections of breast tumours by fluorescence in situ hybridization.

Fluorescence in situ hybridization (FISH) was used to establish the copy number of chromosome 1 in a set of nine breast tumours in which the chromosome had previously been shown to have undergone a variety of rearrangements by loss of heterozygosity studies. In each case, FISH with satellite III DNA from chromosome 1q12 confirmed the results obtained by Southern hybridization. Importantly, in all five cases with rearrangements thought not to involve the centromeric region, FISH showed that the events had not disrupted the gross chromosome structure. This study highlights the potential of using the two techniques together to obtain a clearer picture of both large- and small-scale alterations to chromosomes in solid tumours.

Alleles↗