-
Cell Biochemistry and Biophysics Jan 2012Spectral karyotyping is a novel technique for chromosome analysis that has been developed based on the approach of the fluorescence in situ hybridization technique.... (Review)
Review
Spectral karyotyping is a novel technique for chromosome analysis that has been developed based on the approach of the fluorescence in situ hybridization technique. Spectral karyotyping makes it feasible to diagnose a variety of diseases, because of its technology in painting each of the 24 human chromosomes with different colors. In recent years, it has become possible to adopt the usage of spectral karyotyping for research in general clinical practice, and its usability has attracted particular attention in the diagnosis of different diseases. In this review, we will explain the principle of the spectral karyotyping, as well as its specificity and limitation in detecting the genetic defects within clinical application by presenting two case reports.
Topics: Chromosome Deletion; Chromosome Disorders; Chromosomes; Chromosomes, Human, Pair 13; Humans; Spectral Karyotyping; Trisomy
PubMed: 21948110
DOI: 10.1007/s12013-011-9285-2 -
Ecancermedicalscience 2010
PubMed: 22276033
DOI: 10.3332/ecancer.2010.181 -
Polish Archives of Internal Medicine Aug 2022Throughout the last 50 years, cytogenetic analyses of pretreatment bone marrow and / or blood samples from patients diagnosed with acute myeloid leukemia (AML) revealed... (Review)
Review
Throughout the last 50 years, cytogenetic analyses of pretreatment bone marrow and / or blood samples from patients diagnosed with acute myeloid leukemia (AML) revealed a large number of recurring chromosome aberrations, both structural and numerical. Using standard banding methods and, more recently, molecular cytogenetic techniques, such as fluorescence in situ hybridization, spectral karyotyping, multiplex fluorescence in situ hybridization and comparative genomic hybridization, cytogenetic investigations detect acquired abnormalities that, together with submicroscopic gene mutations and changes in gene expression, strongly influence the clinical features and prognosis of patients with AML. Selected reciprocal translocations and inversions and their molecular counterparts, as well as a number of unbalanced chromosome abnormalities are used, together with bone marrow morphology, immunophenotype, and clinical characteristics, to define separate AML entities in the World Health Organization Classification of Haematolymphoid Tumours. Moreover, cytogenetic findings (and specific gene mutations) are being used in geneticrisk classifications, such as the 2022 European LeukemiaNet classification. Such classifications divide patients into broad prognostic categories: favorable, intermediate, and adverse, which are useful in the management of adults with AML. In this article, I review the present data on recurrent chromosome rearrangements in AML and on correlations between cytogenetic findings and clinical features and treatment outcomes of adult patients diagnosed with AML.
Topics: Adult; Chromosome Aberrations; Comparative Genomic Hybridization; Cytogenetic Analysis; Humans; In Situ Hybridization, Fluorescence; Leukemia, Myeloid, Acute
PubMed: 35848612
DOI: 10.20452/pamw.16300 -
Translational Pediatrics Apr 2014Spectral karyotyping (SKY) is a novel cytogenetic technique, has been developed to unambiguously display and identify all 24 humans chromosomes at one time without a... (Review)
Review
Spectral karyotyping (SKY) is a novel cytogenetic technique, has been developed to unambiguously display and identify all 24 humans chromosomes at one time without a priori knowledge of any abnormalities involved. SKY can discern the aberrations that can't be detected very well by conventional banding technique and Fluorescent in situ hybridization (FISH). So SKY is hyper accurate, hypersensitive, and hyper intuitionist. We will review the elements and application of SKY in leukemia.
PubMed: 26835331
DOI: 10.3978/j.issn.2224-4336.2014.01.02 -
BioTechniques Dec 2015Multispectral karyotyping analyzes all chromosomes in a single cell by labeling them with chromosome-specific probes conjugated to unique combinations of fluorophores....
Multispectral karyotyping analyzes all chromosomes in a single cell by labeling them with chromosome-specific probes conjugated to unique combinations of fluorophores. Currently available multispectral karyotyping systems require the purchase of specialized equipment and reagents. However, conventional laser scanning confocal microscopes that are capable of separating multiple overlapping emission spectra through spectral imaging and linear unmixing can be utilized for classifying chromosomes painted with multicolor probes. Here, we generated multicolor chromosome paints from single-sorted human and mouse chromosomes and developed the Karyotype Identification via Spectral Separation (KISS) analysis package, a set of freely available open source ImageJ tools for spectral unmixing and karyotyping. Chromosome spreads painted with our multispectral probe sets can be imaged on widely available spectral laser scanning confocal microscopes and analyzed using our ImageJ tools. Together, our probes and software enable academic labs with access to a laser-scanning spectral microscope to perform multicolor karyotyping in a cost-effective manner.
Topics: Animals; Cell Line; Chromosomes, Human; Chromosomes, Mammalian; Humans; Karyotyping; Mice; Software
PubMed: 26651513
DOI: 10.2144/000114362 -
Journal of Visualized Experiments : JoVE Feb 2012Conventional method to identify and classify individual chromosomes depends on the unique banding pattern of each chromosome in a specific species being analyzed (1, 2)....
Conventional method to identify and classify individual chromosomes depends on the unique banding pattern of each chromosome in a specific species being analyzed (1, 2). This classical banding technique, however, is not reliable in identifying complex chromosomal aberrations such as those associated with cancer. To overcome the limitations of the banding technique, Spectral Karyotyping (SKY) is introduced to provide much reliable information on chromosome abnormalities. SKY is a multicolor fluorescence in-situ hybridization (FISH) technique to detect metaphase chromosomes with spectral microscope (3, 4). SKY has been proven to be a valuable tool for the cytogenetic analysis of a broad range of chromosome abnormalities associated with a large number of genetic diseases and malignancies (5, 6). SKY involves the use of multicolor fluorescently-labelled DNA probes prepared from the degenerate oligonucleotide primers by PCR. Thus, every chromosome has a unique spectral color after in-situ hybridization with probes, which are differentially labelled with a mixture of fluorescent dyes (Rhodamine, Texas Red, Cy5, FITC and Cy5.5). The probes used for SKY consist of up to 55 chromosome specific probes (7-10). The procedure for SKY involves several steps (Figure 1). SKY requires the availability of cells with high mitotic index from normal or diseased tissue or blood. The chromosomes of a single cell from either a freshly isolated primary cell or a cell line are spread on a glass slide. This chromosome spread is labeled with a different combination of fluorescent dyes specific for each chromosome. For probe detection and image acquisition,the spectral imaging system consists of sagnac interferometer and a CCD camera. This allows measurement of the visible light spectrum emitted from the sample and to acquire a spectral image from individual chromosomes. HiSKY, the software used to analyze the results of the captured images, provides an easy identification of chromosome anomalies. The end result is a metaphase and a karyotype classification image, in which each pair of chromosomes has a distinct color (Figure 2). This allows easy identification of chromosome identities and translocations. For more details, please visit Applied Spectral Imaging website (http://www.spectral-imaging.com/). SKY was recently used for an identification of chromosome segregation defects and chromosome abnormalities in humans and mice with Autosomal Dominant Polycystic Kidney Disease (ADPKD), a genetic disease characterized by dysfunction in primary cilia (11-13). Using this technique, we demonstrated the presence of abnormal chromosome segregation and chromosomal defects in ADPKD patients and mouse models (14). Further analyses using SKY not only allowed us to identify chromosomal number and identity, but also to accurately detect very complex chromosomal aberrations such as chromosome deletions and translocations (Figure 2).
Topics: Animals; Chromosome Aberrations; Female; Humans; Male; Mice; Polycystic Kidney Diseases; Spectral Karyotyping
PubMed: 22330078
DOI: 10.3791/3887 -
Genes Jun 2022Primary human umbilical vein endothelial cells (HUVECs) are consistently the most reliable in vitro model system for studying the inner lining of blood and lymphatic...
Primary human umbilical vein endothelial cells (HUVECs) are consistently the most reliable in vitro model system for studying the inner lining of blood and lymphatic vessels or the endothelium. Primary human cells originate from freshly isolated tissues without genetic manipulation and generally show a modal number of 46 chromosomes with no structural alterations, at least during early passages. We investigated the cytogenetic integrity of HUVECs with conventional (G-banding) and molecular cytogenetic methods (spectral karyotyping (SKY) and fluorescence in situ hybridization (FISH)). Our G-band data shows two X-chromosomes, confirming these HUVECs originate from a female donor. Notably, some cells consistently exhibit an unfamiliar banding pattern on one X chromosome toward the distal end of the long arm (Xq). Our FISH analysis confirms that approximately 50% of these HUVECs have a deletion of the Xq terminal region. SKY analysis indicates that the deleted region is apparently not integrated into any other chromosome. Finally, we demonstrated the presence of a similar Xq deletion in the daughter cell line, EA.hy926, which was generated by fusing HUVECs with A549 (a thioguanine-resistant clone of adenocarcinomic human alveolar basal epithelial cells). These findings will advance comprehension of HUVECs biology and will augment future endothelial studies.
Topics: Cytogenetics; Female; Human Umbilical Vein Endothelial Cells; Humans; In Situ Hybridization, Fluorescence; Karyotyping; Mosaicism
PubMed: 35741774
DOI: 10.3390/genes13061012 -
Archives of Pathology & Laboratory... Aug 2006Sarcomas are rare, numerous in type, and often difficult to definitively classify. Work in the last 2 decades has revealed that a significant subset of sarcomas are... (Review)
Review
CONTEXT
Sarcomas are rare, numerous in type, and often difficult to definitively classify. Work in the last 2 decades has revealed that a significant subset of sarcomas are associated with specific chromosomal translocations producing chimeric (fusion) genes that play a role in the sarcomas' biology and are helpful in their differential diagnosis.
OBJECTIVE
To briefly review the sarcomas associated with specific translocations presenting Ewing sarcoma and synovial sarcoma as archetypes and to further explain how cytogenetic and molecular biologic approaches are being used in the diagnosis of sarcomas.
DATA SOURCES
This work is based on a selected review of the relevant medical and scientific literature and our extensive experience with molecular testing in sarcomas.
CONCLUSIONS
In addition to, and complementing, the traditional diagnostic methods of examination of hematoxylin-eosin stained slides, immunohistochemistry, and sound clinical-pathologic correlation, additional cytogenetic and molecular biologic methods are being increasingly utilized and relied on in sarcoma pathology. These methods include chromosomal karyotyping, fluorescence in-situ hybridization, spectral karyotyping, and polymerase chain reaction- based methods for demonstrating specific chromosomal translocations and fusion genes. Understanding the basis of these methods and their application is critical to better provide accurate and validated specific diagnoses of sarcomas.
Topics: Bone Neoplasms; DNA, Neoplasm; Humans; Karyotyping; Molecular Diagnostic Techniques; Neuroectodermal Tumors, Primitive, Peripheral; Sarcoma; Sarcoma, Ewing; Sarcoma, Synovial; Soft Tissue Neoplasms; Translocation, Genetic
PubMed: 16879024
DOI: 10.5858/2006-130-1199-MDOS -
Journal of Cancer Research and... 2022t(8;21)(q22;q22) is the most frequent recurrent translocation in acute myeloid leukemia (AML) resulting in an in-frame fusion of RUNX1/RUNX1T1 that regulates various... (Review)
Review
Acute myeloid leukemia patients with variant or unusual translocations involving chromosomes 8 and 21 - A comprehensive cytogenetic profiling of three cases with review of literature.
BACKGROUND
t(8;21)(q22;q22) is the most frequent recurrent translocation in acute myeloid leukemia (AML) resulting in an in-frame fusion of RUNX1/RUNX1T1 that regulates various genes involved in the signaling pathways. This leukemogenic alteration is usually associated with a favorable clinical outcome. Variants of t(8;21) can be formed involving a third or fourth chromosome in ~3-4% of t(8;21)-AML. Due to the rarity of variant t(8;21), its clinicopathological features and prognostic significance are still unclear. Here we present three AML cases with cryptic rearrangements of chromosomes 8 and 21 without standard RUNX1/RUNX1T1.
MATERIALS AND METHODS
Conventional karyotyping and fluorescence in situ hybridization and/or spectral karyotyping of the pretreatment bone marrow aspirate of de novo AML patients were performed to delineate chromosomal abnormalities.
RESULTS
We identified three cases with novel variants of t(8;21); der(13)t(8;21;13), isodicentric derivative 8 with chromosome 21[,+idicder(8)(q11.1)t(8;21)(q22;q11.1)] and der(21)t(8;12;21)(q22;q?;q22).
CONCLUSION
AML with t(8;21)(q22;q22);RUNX1-RUNX1T1 forms a distinct WHO subcategory and hence the identification of variants or unusual translocations associated with t(8;21) deserves more attention. Contribution to the variant/ unusual t(8;21) database will further refine the risk stratification and may help to significantly advance the current treatment regimen.
Topics: Chromosomes; Chromosomes, Human, Pair 21; Chromosomes, Human, Pair 8; Core Binding Factor Alpha 2 Subunit; Humans; In Situ Hybridization, Fluorescence; Karyotyping; Leukemia, Myeloid, Acute; Translocation, Genetic
PubMed: 35900542
DOI: 10.4103/jcrt.jcrt_190_21 -
The Journal of Histochemistry and... Aug 2018Aneuploidy seems to play not only a decisive role in embryonal development but also in tumorigenesis where chromosomal and genomic instability reflect a universal...
Aneuploidy seems to play not only a decisive role in embryonal development but also in tumorigenesis where chromosomal and genomic instability reflect a universal feature of malignant tumors. The cost of whole genome sequencing has fallen significantly, but it is still prohibitive for many institutions and clinical settings. No applied, cost-effective, and efficient technique has been introduced yet aiming at research to assess the ploidy status of all 24 different human chromosomes in interphases simultaneously, especially in single cells. Here, we present the selection of human probe DNA and a technique using multistep fluorescence in situ hybridization (FISH) employing four sets of six labeled FISH probes able to delineate all 24 human chromosomes in interphase cells. This full karyotype analysis approach will provide additional diagnostic potential for single cell analysis. The use of spectral imaging (SIm) has enabled the use of up to eight different fluorochrome labels simultaneously. Thus, scoring can be easily assessed by visual inspection, because SIm permits computer-assigned and distinguishable pseudo-colors to each probe during image processing. This enables full karyotype analysis by FISH of single-cell interphase nuclei.
Topics: Aneuploidy; Chromosomes, Artificial, Bacterial; DNA Probes; Humans; Image Processing, Computer-Assisted; In Situ Hybridization, Fluorescence; Interphase; Karyotype; Karyotyping; Male; Plasmids; Single-Cell Analysis
PubMed: 29672206
DOI: 10.1369/0022155418771613