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PubMed1994

Detection of nondisjunction and recombination in meiotic and postmeiotic cells from XYSxr [XY,Tp(Y)1Ct] mice using multicolor fluorescence in situ hybridization.

Current meiotic dogma holds that synapsis is required for recombination and that recombination is required for proper disjunction. The mouse chromosome aberration XYSxr [sex reversal; redesignated XY,Tp(Y)1Ct] appears to challenge this assumption, for although chromosomes X and Y often fail to synapse and recombine, there is no dramatic increase in aneuploid progeny. An explanation of this conundrum might be that X-Y univalent spermatocytes do not survive. The phenotype of sex reversal is generated by the "obligatory" crossover between the X and Y chromosomes, which always occurs proximal to a duplicated copy of the testis-determining gene Sry and transfers one copy from one chromatid of the Y chromosome to one chromatid of the X. Animals that inherit an X chromosome with the Sry gene are chromosomally female but phenotypically male. We have used fluorescence in situ hybridization (FISH) to visualize probes for the X and Y chromosomes and for the Sry sequence and chromosome 8 to track the fate of both recombinant and nonrecombinant chromosomes through metaphases I and II into spermatids and sperm. In the 219 gametes examined by multicolor FISH, the frequency of aneuploid products (XY or "O") was low (3.7%) despite a high frequency (66%) of X-Y separation at metaphase I. In balanced gametes, X and Y recombinant chromosomes slightly exceeded nonrecombinants. Both of these observations support the earlier proposal that asynapsis and nondisjunction in primary spermatocytes lead to their developmental arrest and degeneration.

Aneuploidy↗
PubMedDate not supplied

Bone marrow injury in lithographers exposed to glycol ethers and organic solvents used in multicolor offset and ultraviolet curing printng processes.

Prompted by referral of a printer with aplastic anemia, a study of possible marrow toxicity of workplace substances was undertaken. Dermal and respiratory exposures to dipropylene glycol monomethyl ether, ethylene glycol monoethyl ether, and a range of aliphatic, aromatic and halogenated hydrocarbons used for offset and ultraviolet cured multicolor printing were documented. Evaluation of seven co-workers revealed normal peripheral blood pictures, but bone marrow specimens demonstrated clear patterns of injury in three while the others had nonspecific signs of marrow effect. These changes could not be explained by known risk factors. The authors conclude that further evaluation of possible bone marrow toxicity resulting from exposure to glycol ethers and ultraviolet curing printing processes is warranted. More generally, we have provided data demonstrating that peripheral blood counts may be an insensitive tool for the study of hematologic toxins acting at the bone marrow level.

Adult↗
PubMed2000

Multicolor FISH analysis of chromosomal breaks, duplications, deletions, and numerical abnormalities in the sperm of healthy men.

Transmitted de novo structural chromosomal abnormalities, the majority of which are paternally derived, can lead to abnormal reproductive outcomes as well as genetic diseases in offspring. We developed and validated a new multicolor FISH procedure (sperm ACM, which utilizes DNA probes specific for the alpha [1cen], classical, [1q12], and midi [1p36.3] satellites of chromosome 1) which utilizes DNA probes specific for three regions of chromosome 1 to detect human sperm that carry numerical abnormalities plus two categories of structural aberrations: (1) duplications and deletions of 1pter and 1cen, and (2) chromosomal breaks within the 1cen-1q12 region. In healthy men, the average frequencies of sperm with duplications and deletions were (a) 4.5 +/- 0.5 and 4.1 +/- 1.3 per 10(4) involving 1pter and (b) 0.9 +/- 0.4 and 0.8 +/- 0.3 per 10(4) involving 1cen, respectively. The frequency of sperm exhibiting breaks within the 1cen-1q12 region was 14.1 +/- 1.2 per 10(4). Structural aberrations accounted for 71% of the abnormalities detected by sperm ACM, which was significantly higher than numerical abnormalities (P=2x10-8). Our findings also suggest that, for healthy men, (a) sperm carrying postmeiotic chromosomal breaks appear to be more prevalent than those carrying products of premeiotic or meiotic breakage or rearrangements, (b) the high frequency of chromosome breaks measured after "fertilization" by the hamster-egg cytogenetic method already appear to be present and detectable within human sperm by FISH, and (c) there are nonrandom and donor-specific distributions of breakpoint locations within 1q12 in sperm. FISH facilitates the analysis of much larger numbers of sperm than was possible when the hamster-egg method was used. Therefore, FISH-based procedures for simultaneously detecting chromosomal breaks, rearrangements, and numerical abnormalities in sperm may have widespread applications in human genetics, genetic toxicology, and reproductive medicine.

Animals↗
PubMed1997

Chromosomal alterations affecting the 1cen-1q12 region in buccal mucosal cells of betel quid chewers detected using multicolor fluorescence in situ hybridization.

Epidemiological studies have shown that a high incidence of oral cancers is associated with chewing betel quid. Since chromosomal aberrations are involved in many types of cancers, we investigated whether increased frequencies of chromosomal alterations could be detected in the oral mucosa cells of betel quid chewers as compared to non-chewers. Due to the difficulty in culturing these epithelial cells, we used multicolor FISH with adjacent DNA probes to detect hyperdiploidy and breakage/exchanges affecting the 1cen-q12 region in interphase cells. Buccal mucosa cells from 19 male betel quid chewers and 23 non-chewers were hybridized and 1000 cells per donor were evaluated. A highly significant increase in the frequency of breakage affecting 1cen-1q12 region was observed in the mucosa cells of the chewers as compared to the non-chewers. A good correlation was also seen between breakage and duration of chewing. A modest increase in hyperdiploidy for chromosome 1 was also observed among chewers who had used betel quid for many years. These results indicate that this FISH approach can be useful for human biomonitoring, particularly for detecting alterations in non-dividing cells.

Areca↗
PubMed1992

Multicolor fluorescence in situ hybridization for the simultaneous detection of probe sets for chromosomes 13, 18, 21, X and Y in uncultured amniotic fluid cells.

The most frequent aneuploidies in newborns involve the autosomes 13, 18 and 21 as well as both sex chromosomes. Fluorescence in situ hybridization readily allows the detection of numerical chromosomal aberrations throughout all stages of the cell cycle. Using a multicolor fluorescence in situ hybridization approach based on combinatorial probe labeling and digital imaging microscopy we demonstrate the simultaneous visualization of probe sets specific for chromosomes 13, 18, 21, X and Y. This approach enables one to evaluate aberrations of multiple chromosomes in a single hybridization experiment using metaphase chromosomes and interphase nuclei from a variety of cell types, including lymphocytes and amniocytes.

Amniotic Fluid↗
PubMed2003

The multicolored Asian lady beetle, Harmonia axyridis: a review of its biology, uses in biological control, and non-target impacts.

Throughout the last century, the multicolored Asian lady beetle, Harmonia axyridis (Pallas) has been studied quite extensively, with topics ranging from genetics and evolution to population dynamics and applied biological control being covered. Much of the early work on H. axyridis was conducted in the native Asian range. From the 1980's to the present, numerous European and North American studies have added to the body of literature on H. axyridis. H. axyridis has recently gained attention in North America both as a biological control agent and as a pest. This literature review was compiled for two reasons. First, to assist other researchers as a reference, summarizing most of the voluminous body of literature on H. axyridis pertaining to its biology, life history, uses in biological control, and potential non-target impacts. Secondly, to be a case study on the impacts of an exotic generalist predator.

Animals↗
PubMed2001

Chromosomal composition of micronuclei in mouse NIH 3T3 cells treated with acrylamide, extract of Tripterygium hypoglaucum (level) hutch, mitomycin C and colchicine, detected by multicolor FISH with centromeric and telomeric DNA probes.

The chromosomal composition of micronuclei (MN) induced by the model mutagens mitomycin (MMC) and colchicine (COL) as well as by acrylamide (AA) and the traditional Chinese medicine Tripterygium hypoglaucum (level) hutch (THH) in NIH 3T3 cells was analyzed by multicolor fluorescence in situ hybridization (FISH) using DNA probes for the centromere repeated minor satellite DNA and the telomeric hexamer repeat (TTAGGG). The majority of MN (78.6%) from treatment with MMC (0.1 microg/ml) did not show centromeric signals, reflecting the clastogenic action of MMC. Following treatment with COL (0.1 microg/ml), 74.5% of the MN showed centromeric signals and several telomeric signals, indicating that MN induced by this well-known aneugen were mainly composed of whole chromosomes. After treatment with AA (100, 200 and 400 microg/ml) both MN containing whole chromosomes and MN containing acentric fragments were found to increase in a dose-dependent manner, demonstrating that AA is not only a clastogen but also an aneugen. THH induced a high frequency of MN harboring whole chromosomes at all concentrations tested (5, 10 and 20 microl/ml) and produced a dose-dependent increase in fragment-containing MN, indicating that THH has both aneugenic and clastogenic potential.

3T3 Cells↗
PubMed1994

Detection of hydroquinone-induced nonrandom breakage in the centromeric heterochromatin of mouse bone marrow cells using multicolor fluorescence in situ hybridization with the mouse major and minor satellite probes.

Fluorescence in situ hybridization with a mouse major satellite probe and CREST staining were used to characterize the origin of micronuclei occurring in mouse bone marrow erythrocytes following administration of the benzene metabolite hydroquinone. Hydroquinone was administered to male CD-1 mice by i.p. injection on three consecutive days and the bone marrow cells were harvested 24 h later. A pronounced difference in the results was observed using the two approaches: 63% of the micronuclei induced by hydroquinone labeled with the major satellite probe whereas only 28% labeled with the CREST antibody. To determine whether the observed difference was due to a disruption of the kinetochore or a result of breakage within centromeric heterochromatin, we developed a tandem label multicolor hybridization assay, which requires the presence of both the mouse major and minor satellite probes in a micronucleus for a classification of chromosomal loss. The minor probe targets a centromeric region physically linked to the short arm of mouse chromosomes, whereas the major probe hybridizes to the centromeric heterochromatin adjacent to the long arm. Using this approach, 29% of the micronuclei induced by hydroquinone hybridized with both the major and minor satellite probes, indicating chromosome loss; an additional 37% labeled with only the major satellite probe indicating breakage within the centromeric heterochromatin. Although the region targeted by the major satellite probe comprises only 5-10% of the mouse genome, these major-probe containing micronuclei represent 53% of the micronuclei formed as the result of chromosome breakage.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2005

Detection of single DNA molecules by multicolor quantum-dot end-labeling.

Observation of DNA-protein interactions by single molecule fluorescence microscopy is usually performed by using fluorescent DNA binding agents. However, such dyes have been shown to induce cleavage of the DNA molecule and perturb its interactions with proteins. A new method for the detection of surface-attached DNA molecules by fluorescence microscopy is introduced in this paper. Biotin- and/or digoxigenin-modified DNA fragments are covalently linked at both extremities of a DNA molecule via sequence-specific hybridization and ligation. After the modified DNA molecules have been stretched on a glass surface, their ends are visualized by multicolor fluorescence microscopy using conjugated quantum dots (QD). We demonstrate that under carefully selected conditions, the position and orientation of individual DNA molecules can be inferred with good efficiency from the QD fluorescence signals alone. This is achieved by selecting QD pairs that have the distance and direction expected for the combed DNA molecules. Direct observation of single DNA molecules in the absence of DNA staining agent opens new possibilities in the fundamental study of DNA-protein interactions. This work also documents new possibilities regarding the use of QD for nucleic acid detection and analysis.

Biotinylation↗
PubMed2005

A set of multicolored Photinus pyralis luciferase mutants for in vivo bioluminescence applications.

Error-prone PCR was used to isolate Photinus pyralis luciferase mutants producing bright light in the red-orange region of the spectrum. All mutations were clustered in the beta5-alpha10-beta6 region of N-terminal subdomain B and appear to affect bioluminescence color by modulating the position of the Ser314-Leu319 mobile loop with respect to the putative active site. Two red variants (Q283R and S284G) and one orange mutant (S293P) contained a single substitution. Although the remaining orange variant contained two mutations, L287I mainly contributed to the color change. Emission spectra collected on whole cells at pH 7.0 revealed that while a single peak of lambdamax approximately 605 nm accounts for red light production by the Q283R and S284G variants, orange light results from the contribution of two peaks of lambdamax approximately 560 and 600 nm. All spectra underwent a red-shift when cells were assayed under acidic conditions, whereas a blue-shift was observed at pH 8.0, indicating that the internal pH of Escherichia coli is close to the external pH shortly after imposition of acid or alkaline stress. In addition, changes in assay pH led to bimodal emission spectra, lending support to the idea that bioluminescence color is determined by the relative contribution of yellow-green and red-orange peaks. The set of multicolored luciferase mutants described here may prove useful for a variety of applications including biosensing, pH monitoring, and tissue and animal imaging.

Animals↗
PubMed2003

Multicolor chromosomal bar coding characterizes a de novo interstitial deletion (5)(q33.3q35.2) in a child with multiple congenital malformations.

We describe a boy with multiple congenital anomalies including a complex heart defect, club feet, adducted thumbs, and facial dysmorphic features. He died at the age of 2 months following cardiac surgery. G-banding analysis identified an abnormal chromosome 5q suspected to be an interstitial deletion (5)(q33q35). Breakpoints of the deleted segment were confirmed as del(5)(q33.3q35) by multicolor fluorescence in situ hybridization (FISH) using two sets of combinatorially labeled band specific YAC clones. Findings are discussed in view of previously published cases.

Abnormalities, Multiple↗
PubMedDate not supplied

Multicolor flow-cytometric, morphologic, and clonogenic analysis of marrow CD10-positive cells in children with leukemia in remission or nonmalignant diseases.

PURPOSE: To characterize nonleukemic CD10-positive cells in the marrows of children with leukemia in remission or benign conditions. PATIENTS AND METHODS: Seventeen children with acute leukemia in complete remission, 12 with solid tumors, and 17 with benign blood diseases were included in this study. Bone marrow cells were analyzed by multicolor flow-cytometry and polymerase chain reaction (PCR) to detect immunoglobulin gene rearrangement. The CD10-positive cells were purely sorted and examined by light microscopy and single cell hemopoietic progenitor assay. RESULTS: In patients with acute leukemia, CD10-positive cells were present in higher proportion after completion of therapy than during chemotherapy. They were also higher in the patients of preschool age than in the older age group with benign blood diseases and solid tumors. These CD10-positive cells were morphologically compatible with immature lymphocytes but some blast-like cells also occurred in this population. Most CD10-positive cells coexpressed CD19 and HLA-DR, although only 10 to 30% coexpressed CD20 and CD34. Although some CD10-positive cells expressed CD34, they did not make any colonies. PCR analysis did not show monoclonal bands in CD10-positive bone marrow cells in any patients in remission. CONCLUSION: Marrow CD10-positive cells possess immature B-lymphocyte phenotype and are present in higher proportion in the marrows of children with acute leukemia in continuous complete remission after completion of therapy and children of preschool age than school-age children with benign diseases or solid tumors without marrow involvement. The clonality of these cells was excluded by PCR, which is a distinct point from CD10-positive ALL blasts.

Bone Marrow Cells↗
PubMedDate not supplied

Utility of multicolor fluorescent in situ hybridization in clinical cytogenetics.

PURPOSE: Multicolor FISH (M-FISH) was introduced in 1996 to scan all 24 chromosomes in different fluorescent colors by use of a specific filter set and computer software. However, the clinical utility of M-FISH has been limited because of the lack of commercial availability of reagents and hardware. We have evaluated M-FISH for identification of markers, derivative chromosomes, and complex karyotypes. METHODS: We present our findings based on a representative sample of one normal and six abnormal cases from a variety of tissue types. The results of M-FISH were confirmed by other well-established FISH probes. RESULTS: M-FISH analyses were successful in all six cases. The derivative chromosomes, ring, and a complex karyotype were resolved. CONCLUSIONS: We find M-FISH to be an invaluable tool for a high degree of accuracy and efficiency for chromosome identification. The limitations similar to spectral karyotyping system (SKY) include the inability to detect intrachromosomal anomalies, abnormalities involving the p-arms of acrocentrics and areas rich in highly repetitive DNA. In addition, there are some concerns of misinterpretation due to overlap of fluorophore combinations of different chromosomes, especially for subtle insertional translocations.

Adult↗
PubMedDate not supplied

Multicolor fluorescence-based approaches for imaging cytokine-induced alterations in the neovascularization, growth, metastasis, and apoptosis of murine neuroblastoma tumors.

Neuroblastoma is one of the most common solid tumors in children. The prognosis of patients with advanced neuroblastoma is poor overall despite standard therapeutic modalities and has stimulated substantial interest in the potential role for biologics such as immunotherapeutic and/or antiangiogenic agents for the treatment of neuroblastoma. To facilitate preclinical investigation of the efficacy and mechanisms of action of new biologic agents for the treatment of neuroblastoma, a comprehensive panel of disease-specific fluorescence-based model systems has been developed by our group to image the growth, neovascularization, metastasis, and apoptosis of neuroblastoma tumors. These model systems use fluorescent proteins to monitor cytokine-induced alterations in the growth and metastasis of neuroblastoma and allow for monitoring and/or quantitation of even minimal residual disease that is localized to visceral organ sites such as the liver, lung, and/or bone marrow. Further, based on the differential spectra of red fluorescent protein, green fluorescent protein (GFP), and agents such as 4'-6-diamidino-2-phenylindole (DAPI) (blue) and fluorescein isothiocyanate-dextran (green), multicolor systems have now been established by our group that allow for combined assessment of parameters, including the macroscopic relation of tumors to their associated vasculature and, within tissue sections, simultaneous quantitation of tumor neovascularization and evaluation of therapy-induced apoptosis within the tumor and vascular endothelial compartments. Further, by engineering cells to express specific mediators of apoptosis that have been linked to GFP (ie, BID-EGFP), these systems can also be used to dissect mechanisms by which neuroblastoma cells are induced to undergo apoptosis in vitro as well as in vivo. Collectively, these model systems provide important tools for investigation of the biology of neuroblastoma tumors and evaluation of mechanisms that mediate the regression of these tumors in response to novel therapeutic agents, including cytokines such as interleukin-12.

Angiogenesis Inhibitors↗
PubMed2003

Multicolor karyotyping and clinicopathological analysis of three intravascular lymphoma cases.

Intravascular lymphoma (IVL) is a rare neoplastic disease characterized by the presence of large malignant lymphoid cells in small vessels. It is often diagnosed at autopsy. Clinical manifestations are typically neurologic and dermatologic. Karyotypic abnormalities have been described in a small number of cases and have revealed complex alterations in the majority of cases. We have identified three cases of IVL with varied clinicopathological findings. Karyotypic analysis was undertaken by standard G-banding and supplemented by multi-colored karyotyping (M-FISH) to decipher the chromosomal content of marker chromosomes and undefined additions. M-FISH clarified the chromosomal abnormalities in two cases and unveiled cryptic translocations der(10)t(10;22), der(17)t(17;22), and balanced t(11;14). Comparison with previously published karyotypes revealed prominent involvement of chromosomes 1, 3, 6, 11, 14, and 18, similar to the pattern of clonal evolution in other B-cell lymphomas. The most frequent alterations seen were -6 or 6q- and +18 or dup(18q), with a minimally deleted region located at 6q21-q23 and a commonly amplified region located at 18q13-q23, respectively. Few differences between the classical and Asian variant of this disease were apparent at the karyotypic level. Cytogenetic analysis of additional cases supplemented by multicolor karyotyping may help identify the full spectrum of genetic alterations associated with IVL and assist in the delineation of the critical mutations associated with initiation and progression of this disease.

Aged↗
PubMed2005

Multicolor vortex solitons in two-dimensional photonic lattices.

We report on the existence and stability of multicolor lattice vortex solitons constituted by coupled fundamental frequency and second-harmonic waves in optical lattices in quadratic nonlinear media. It is shown that the solitons are stable almost in the entire domain of their existence, and that the instability domain decreases with the increase of the lattice depth. We also show the generation of the solitons, and the feasibility of the concept of lattice soliton algebra.

Journal Article↗
PubMed2001

Semiconductor quantum dot: a quantum light source of multicolor photons with tunable statistics.

We investigate the intensity correlation properties of single photons emitted from an optically excited single semiconductor quantum dot. The second order temporal coherence function of the photons emitted at various wavelengths is measured as a function of the excitation power. We show experimentally and theoretically that a quantum dot is not only a source of nonclassically correlated monochromatic photons but is also a source of multicolor photons with tunable correlation properties. We found that the emitted photon statistics can be varied by the excitation rate from a sub-Poissonian one, where the photons are temporally antibunched, to super-Poissonian, where they are temporally bunched.

Journal Article↗
PubMed2002

Solution of the multi-Reggeon compound state problem in multicolor QCD.

We study the properties of the color-singlet compound states of Reggeized gluons in multicolor QCD using their relation with non-compact two-dimensional Heisenberg spin magnets. Applying the methods of integrable models, we calculate their spectrum and discuss the application of the obtained results to high-energy asymptotics of the scattering amplitudes in perturbative QCD.

Journal Article↗