Multicolor Karyotyping Research Articles

A Central Role of Telomere Dysfunction in the Formation of a Unique Translocation within the Sub-Telomere Region Resulting in Duplication and Partial Trisomy

Telomeres play a major role in maintaining genome stability and integrity. Putative involvement of telomere dysfunction in the formation of various types of chromosomal aberrations is an area of active research. Here, we report a case of a six-month-old boy with a chromosomal gain encompassing the 11q22.3q25 region identified by SNP array analysis. The size of the duplication is 26.7 Mb and contains 170 genes (OMIM). The duplication results in partial trisomy of the region in question with clinical consequences, including bilateral renal dysplasia, delayed development, and a heart defect. Moreover, the karyotype determined by R-banding and chromosome painting as well as by hybridization with specific sub-telomere probes revealed the presence of an unbalanced t(9;11)(p24;q22.3) translocation with a unique breakpoint involving the sub-telomere region of the short arm of chromosome 9. The karyotypes of the parents were normal. Telomere integrity in circulating lymphocytes from the child and from his parents was assessed using an automated high-throughput method based on fluorescence in situ hybridization (FISH) with telomere- and centromere-specific PNA probes followed by M-FISH multicolor karyotyping. Very short telomeres, as well as an increased frequency of telomere loss and formation of telomere doublets, were detected in the child’s cells. Interestingly, similar telomere profiles were found in the circulating lymphocytes of the father. Moreover, an assessment of clonal telomere aberrations identified chromosomes 9 and 11 with particularly high frequencies of such aberrations. These findings strongly suggest that telomere dysfunction plays a central role in the formation of this specific unbalanced chromosome rearrangement via chromosome end-to-end fusion and breakage–fusion–bridge cycles.

ALK-positive diffuse large B-cell lymphoma is associated with Clathrin-ALK rearrangements: report of 6 cases

AbstractExpression of ALK protein by lymphoid cells and the description of variant anaplastic lymphoma kinase (ALK) translocations have typically been restricted to cases of T-cell and null anaplastic large-cell lymphoma (ALCL). All such cases result from a novel fusion created by the ALK gene on chromosome 2p23 and NPM on 5q35 or other variant translocation partners. A rare variant of diffuse large B-cell lymphoma (DLBCL), originally described in 1997, was thought to overexpress full-length ALK in contrast to a chimeric protein characteristic of ALCL. However, full-length ALK protein lacks tyrosine kinase activity and thus the mechanism of oncogenesis has remained elusive. We describe 6 cases of ALK+ DLBCL characterized by a simple or complex t(2;17)(p23;q23) involving the clathrin gene (CLTC) at chromosome band 17q23 and the ALK gene at chromosome band 2p23. All cases were studied using fluorescence in situ hybridization (FISH), complemented in one case with standard cytogenetic analysis, multicolor karyotyping (M-FISH), and reverse transcriptase-polymerase chain reaction. These results clearly demonstrate that most cases of ALK+ DLBCL share the same mechanism of deregulated ALK expression. Moreover, these results demonstrate the presence of CLTC-ALK fusions in these tumors and extend the list of diseases associated with this genetic abnormality to include classical T-cell or null ALCL, ALK+ DLBCL, and inflammatory myofibroblastic tumors. (Blood. 2003;102:2568-2573)

Molecular cytogenetic analysis of genome-specific repetitive elements in Citrus clementina Hort. Ex Tan. and its taxonomic implications

BackgroundClementine mandarin (Citrus clementina Hort. ex Tan.) is one of the most famous and widely grown citrus cultivars worldwide. Variations in relation to the composition and distribution of repetitive DNA sequences that dominate greatly in eukaryote genomes are considered to be species-, genome-, or even chromosome-specific. Repetitive DNA-based fluorescence in situ hybridization (FISH) is a powerful tool for molecular cytogenetic study. However, to date few studies have involved in the repetitive elements and cytogenetic karyotype of Clementine.ResultsA graph-based similarity sequence read clustering methodology was performed to analyze the repetitive DNA families in the Clementine genome. The bioinformatics analysis showed that repetitive DNAs constitute 41.95% of the Clementine genome, and the majority of repetitive elements are retrotransposons and satellite DNAs. Sequential multicolor FISH using a probe mix that contained CL17, four satellite DNAs, two rDNAs and an oligonucleotide of (TTTAGGG)3 was performed with Clementine somatic metaphase chromosomes. An integrated karyotype of Clementine was established based on unequivocal and reproducible chromosome discriminations. The distribution patterns of these probes in several Citrus, Poncirus and Fortunella species were summarized through extensive FISH analyses. Polymorphism and heterozygosity were commonly observed in the three genera. Some asymmetrical FISH loci in Clementine were in agreement with its hybrid origin.ConclusionsThe composition and abundance of repetitive elements in the Clementine genome were reanalyzed. Multicolor FISH-based karyotyping provided direct visual proof of the heterozygous nature of Clementine chromosomes with conspicuous asymmetrical FISH hybridization signals. We detected some similar and variable distribution patterns of repetitive DNAs in Citrus, Poncirus, and Fortunella, which revealed notable conservation among these genera, as well as obvious polymorphism and heterozygosity, indicating the potential utility of these repetitive element markers for the study of taxonomic, phylogenetic and evolutionary relationships in the future.

Multicolor Karyotyping and Fluorescence In Situ Hybridization-Banding (MCB/mBAND).

Multicolor fluorescence in situ hybridization (mFISH) approaches are routine applications in tumor as well as clinical cytogenetics nowadays. The first approach when thinking about mFISH is multicolor karyotyping using human whole chromosome paints as probes; this can be achieved by narrow-band filter-based multiplex-FISH (M-FISH) or interferometer/spectroscopy-based spectral karyotyping (SKY). Besides, various FISH-based banding approaches were reported in the literature, including multicolor banding (MCB/mBAND) the latter being evaluated by narrow-band filters, and using specific software. Here, we describe the combined application of multicolor karyotyping and MCB/mBAND for the characterization of simple and complex acquired chromosomal changes in cancer cytogenetics.

Karyotyping human and mouse cells using probes from single-sorted chromosomes and open source software.

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.

Molecular cytogenetics : an indispensable tool for cancer diagnosis

Cytogenetic aberrations may escape detection or recognition in traditional karyotyping. The past decade has seen an explosion of methodological advances in molecular cytogenetics technology. These cytogenetics techniques add color to the black and white world of conventional banding. Fluorescence in-situ hybridization (FISH) study has emerged as an indispensable tool for both basic and clinical research, as well as diagnostics, in leukemia and cancers. FISH can be used to identify chromosomal abnormalities through fluorescent labeled DNA probes that target specific DNA sequences. Subsequently, FISH-based tests such as multicolor karyotyping, comparative genomic hybridization (CGH) and array CGH have been used in emerging clinical applications as they enable resolution of complex karyotypic aberrations and whole global scanning of genomic imbalances. More recently, crossspecies array CGH analysis has also been employed in cancer gene identification. The clinical impact of FISH is pivotal, especially in the diagnosis, prognosis and treatment decisions for hematological diseases, all of which facilitate the practice of personalized medicine. This review summarizes the methodology and current utilization of these FISH techniques in unraveling chromosomal changes and highlights how the field is moving away from conventional methods towards molecular cytogenetics approaches. In addition, the potential of the more recently developed FISH tests in contributing information to genetic abnormalities is illustrated.

A novel unbalanced de novo translocation der(5)t(4;5)(q26;q21.1) in adult T-cell precursor lymphoblastic leukemia

We here describe a novel unbalanced de novo translocation der(5)t(4;5)(q26;q21.1) in a 39-year-old male diagnosed with acute T-cell lymphoblastic leukemia. Bone marrow (BM) was massively infiltrated with 85 % highly proliferative polymorphic T-cell precursors. Immunologically, the malignant cells stained positive for CD7, CD34, intracytoplasmic CD3+, TdT + and negative for CD3 and CD5. G-banded chromosome analysis of BM cells showed the normal karyotype 46,XY[25] whereas BAC-based aCGH analysis revealed partial gain of 4q and partial loss of 5q. Multicolor karyotyping confirmed the presence of an unbalanced der(5)t(4;5) as the sole structural abnormality. Subsequent high-resolution oligonucleotide-based aCGH analysis showed that the der(5)t(4;5)(q26;q21.1) resulted in partial trisomy of 4q26qter (117,719,015-190,613,014) and partial monosomy of 5q21.1qter (100,425,442-180,857,866) and that there was no indication of any gene disruptions resulting from the breakages. Interphase FISH analysis using BAC-based specific probes for 4q26 and 5q21.1 confirmed the breakpoints and revealed approximately 80 % abnormal cells accordingly. At 4q26 the MIR1973 gene is located centromeric to the breakpoint in the copy number neutral region and the TRAM1L1 gene is located within the gained region. At 5q21.1 the genes ST8SIA4 and MIR548p are located centromeric to the breakpoint and no known genes up to approximately 1 Mb telomeric to the breakpoint in the copy number loss region. Interestingly, only the gene ST8SIA4 at 5q21.1 have been implicated in T-cell regulation as it encodes one of the key enzymes for polysialysation of surface proteins on dendritic cells which are important regulators for T-cell proliferation. The der(5)t(4;5) is thought to play a crucial role in the pathogenesis of acute T-ALL due to either gain of 4q, the loss of 5q, or deregulation of genes in proximity to the breakpoints.

Combining M-FISH and Quantum Dot technology for fast chromosomal assignment of transgenic insertions

BackgroundPhysical mapping of transgenic insertions by Fluorescence in situ Hybridization (FISH) is a reliable and cost-effective technique. Chromosomal assignment is commonly achieved either by concurrent G-banding or by a multi-color FISH approach consisting of iteratively co-hybridizing the transgenic sequence of interest with one or more chromosome-specific probes at a time, until the location of the transgenic insertion is identified.ResultsHere we report a technical development for fast chromosomal assignment of transgenic insertions at the single cell level in mouse and rat models. This comprises a simplified 'single denaturation mixed hybridization' procedure that combines multi-color karyotyping by Multiplex FISH (M-FISH), for simultaneous and unambiguous identification of all chromosomes at once, and the use of a Quantum Dot (QD) conjugate for the transgene detection.ConclusionsAlthough the exploitation of the unique optical properties of QD nanocrystals, such as photo-stability and brightness, to improve FISH performance generally has been previously investigated, to our knowledge this is the first report of a purpose-designed molecular cytogenetic protocol in which the combined use of QDs and standard organic fluorophores is specifically tailored to assist gene transfer technology.

Small blue round cell tumor of the interosseous membrane bearing a t(2;22)(q34;q12)/EWS-CREB1 translocation: a case report

BackgroundThe group of small blue round cell tumors encompasses a heterogeneous group of neoplasms characterized by primitive appearing round cells with few distinguishing histologic features.ResultsWe report the case of a small blue round cell tumor with an EWS gene rearrangement detected by fluorescent in situ hybridization (FISH) analysis that mimicked Ewing sarcoma, but with unusual histology and immunohistochemical features. Multi-color karyotyping identified the presence of a t(2;22)(q34;q12) that was initially expected to represent a variant EWSR1-FEV translocation. After an extensive workup, the lesion is considered to represent a clear cell sarcoma harboring an EWSR1-CREB1 fusion transcript.ConclusionsThis case appears to represent a rare variant of clear cell sarcoma arising in peripheral soft tissues with unusual histology and unique immunophenotype. In this circumstance, FISH for all EWSR1 translocation partners or RT- PCR for a spectrum of possible transcript variants is critically important for diagnosis, since cytogenetic analysis or clinical FISH assay using only commercial EWSR1 probes will be misleading.

Concurrent gain of 17q and the MYC oncogene in a medullomyoblastoma

Medullomyoblastoma (MMB) is a rare cerebellar childhood tumor containing both myoblastic and primitive neuroectodermal components. Similar to the scenario in classical medulloblastoma, which contains the primitive neuroectodermal component only, gain of sequences from the long arm of chromosome 17 (17q) and gain of the MYC gene in 8q have been implicated in the pathogenesis of MMB. Because karyotypic analysis has not previously been performed for MMB, the mechanisms behind genomic imbalances in this tumor have remained unknown. Several other central aspects of this tumor, such as histocytogenetic origin, clinical characteristics, tumor behavior and prognosis, also remain unknown. We here report neuropathological and cytogenetic features of an MMB in a 3-year-old boy. Chromosome banding analysis and multicolor karyotyping revealed a hyperdiploid karyotype including an unbalanced 1; 17 translocation and isochromosome 17q formation, both leading to gain of 17q. There were also two extra copies of chromosome 8, leading to gain of the MYC oncogene, trisomies 5 and 13, and monosomy 9. Clonal chromosome changes were present in both the myoblastic and neuroectodermal components. Our findings support the notion that MMB and classical medulloblastoma arise through similar genetic mechanisms and that the two main tissue components in MMB are clonally related.

Does parosteal liposarcoma differ from other atypical lipomatous tumors/well-differentiated liposarcomas? A molecular cytogenetic study using combined multicolor COBRA-FISH karyotyping and array-based comparative genomic hybridization

Parosteal adipocytic tumors of the bone are extremely rare. As a result, (cyto-) genetic data on this entity are essentially lacking. In the literature there is debate as to whether these lesions should be classified according to the criteria used in soft-tissue tumor pathology, or if they should be considered a separate bone tumor entity. Here we present a 68-year-old male patient with a tumor in his right upper leg diagnosed as parosteal atypical lipomatous tumor/well-differentiated liposarcoma (ALT/WDLS) on the basis of clinico-radiologic and pathologic findings. Molecular cytogenetic investigations using combined binary ratio labeling fluorescence in situ hybridization and array comparative genomic hybridization showed abnormalities, which are in accordance with the histologic appearance of an atypical lipomatous tumor/well-differentiated liposarcoma. Therefore, on the basis of these molecular cytogenetic investigations, we conclude that parosteal liposarcoma is not a separate entity but should be categorized within the spectrum of soft-tissue ALT/WDLS.

Prenatal diagnosis of nonmosaic tetrasomy 9p by microdissection and FISH: case report

Marker chromosome(mar)is a structurally abnormal chromosome that can not be identified with conventional cytogenetic techniques(ISCN,2005).The incidence of supernumerary marker chromosomes(SMCs) found at prenatal diagnosis varies from 0.4/1000 to 1.5/1000.Their genetic effects may range from harmless to detrimental,and in de novo cases,the rate of phenotypic abnormalities is statistically increased. Therefore,identification of SMCs has become imperative in prenatal diagnosis. A variety of molecular cytogenetic approaches have been developed to identify SMCs,including fluorescence in situ hybridization(FISH),microdissection and reverse FISH(micro-FISH),multicolor karyotyping techniques, centromere-specific multicolor FISH assays,and AcroM-FISH technique,etc.Among these approaches, micro-FISH is the most straightforward method that can accurately detect the origin of SMC and the location of chromosome breakpoint.Herein,we diagnosed a case of tetrasomy 9p by classical cytogenetic methods combined with chromosome microdissection and FISH.

Cryptic terminal chromosome rearrangements in colorectal carcinoma cell lines detected by subtelomeric FISH analysis

Epithelial tumour karyotypes are often difficult to study by standard cytogenetic methods because of poor chromosome preparation quality and the high complexity of their genomic rearrangements. Subtelomeric fluorescence in situ hybridisation (FISH) has proved to be a useful method for detecting cryptic constitutional chromosomal rearrangements but little is known about its usefulness for tumour cytogenetic analysis. Using a combination of chromosome banding, multicolour karyotyping and subtelomeric FISH, five colorectal cancer cell lines were characterised. The resulting data were compared to results from previous studies by comparative genomic hybridisation and spectral karyotyping or multicolour FISH. Subtelomeric FISH made it possible to resolve several highly complex chromosome rearrangements, many of which had not been detected or were incompletely characterised by the other methods. In particular, previously undetected terminal imbalances were found in the two cell lines not showing microsatellite instability.

Wavelet-Based Compression of M-FISH Images

Multiplex fluorescence in situ hybridization (M-FISH) is a recently developed technology that enables multi-color chromosome karyotyping for molecular cytogenetic analysis. Each M-FISH image set consists of a number of aligned images of the same chromosome specimen captured at different optical wavelength. This paper presents embedded M-FISH image coding (EMIC), where the foreground objects/chromosomes and the background objects/images are coded separately. We first apply critically sampled integer wavelet transforms to both the foreground and the background. We then use object-based bit-plane coding to compress each object and generate separate embedded bitstreams that allow continuous lossy-to-lossless compression of the foreground and the background. For efficient arithmetic coding of bit planes, we propose a method of designing an optimal context model that specifically exploits the statistical characteristics of M-FISH images in the wavelet domain. Our experiments show that EMIC achieves nearly twice as much compression as Lempel-Ziv-Welch coding. EMIC also performs much better than JPEG-LS and JPEG-2000 for lossless coding. The lossy performance of EMIC is significantly better than that of coding each M-FISH image with JPEG-2000.

FISHing — Current Status and Future Prospects for Improved Management of Leukemia

It is now mandatory to perform cytogenetics (CG) study in all new cases of leukemia owing to its usefulness in disease diagnosis, classification and prognostication. The technique of fluorescence in-situ hybridization (FISH), applicable to both interphase and metaphase cells on a variety of samples has proved to be useful in supplementing convention CG, especially in the detection of clinically significant changes when CG is unsuccessful or when genetic abnormalities are cryptic. With improvements in the test sensitivity, FISH can be applied in monitoring molecular response to treatment and detection of residual disease, especially in the setting of monitoring for BCR-ABL in chronic myeloid leukemia patients following imatinib therapy. Newer molecular CG tests such as multicolor karyotyping and comparative genomic hybridization have a clinical potential as they enable resolution of complex karyotypic aberrations and global scanning of genomic imbalances, respectively. In the horizon, important information gained from gene expression profile studies will be useful for the design of tests, molecular based or otherwise, that are suited for routine diagnostic service. However, CG and molecular data must also be integrated with clinical and pathological findings before a definitive diagnosis is made.

Multicolour fluorescence in situ hybridization analysis of t(14;18)‐positive follicular lymphoma and correlation with gene expression data and clinical outcome

In order fully to identify secondary chromosomal alterations, such as duplications, additions and marker chromosomes that remained unresolved by G banding, 60 cases of t(14;18)-positive follicular lymphoma (FL) were analysed by multicolour karyotyping techniques [multicolour fluorescence in situ hybridization (MFISH)/multicolour banding for chromosome 1 (MBAND1)]. A total of 165 additional structural chromosomal aberrations were delineated. An increased frequency of chromosomal gains involving X, 1q, 2, 3q27-q29, 5, 6p11-p21, 7, 8, 11, 12, 14q32, 17q, 18 and 21 and deletions of 1p36, 3q28-q29, 6q, 10q22-q24 and 17p11-p13 was revealed by the MFISH/MBAND1 analysis. Balanced translocations other than t(14;18) were uncommon, whereas unbalanced translocations were numerous. Deletion of 1p36 and duplication of 1p33-p35, 1p12-p21 and 1q21-q41 were regularly involved in chromosome 1 alterations, seen in 53% of the cases. A strong correlation was demonstrated between gains of individual chromosomal bands and increased gene expression, including 1q22/MNDA, 6p21/CDKN1A, 12q13-q14/SAS, 17q23/ZNF161, 18q21/BCL2 and Xq13/IL2RG. Unfavourable overall survival was associated with del(1)(p36) and dup(18q). These data support the notion that translocation events are primarily responsible for FL disease initiation, whereas the unbalanced chromosomal gains and losses that mirror the gene expression patterns characterize clonal evolution and disease progression, and thus provide further insights into the biology of FL.

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.

Advances in molecular cytogenetics for the evaluation of mental retardation.

Recent years have witnessed rapid advances in molecular cytogenetics and its impact in studying mental retardation (MR). We review new molecular cytogenetic methods, including interphase fluorescence in situ hyrbridization (FISH), comparative genomic hybridization (CGH), multicolor karyotyping, telomere FISH, primed in situ labeling (PRINS), genotyping, microdissection, and microarray for the evaluation of MR. These new methods are very useful in two major aspects: further characterization of chromosome abnormalities as detected with routine banding analysis, including additions, duplications, deletions, translocations, markers, or complex aberrations; and screening for "hidden" chromosome aberrations in patients with an apparently normal karyotype. These new methods have great diagnostic potential in prenatal, postnatal, and preimplantational settings. Although powerful, at this point, they are primarily research tools in nature. It is essential that these new methods be used in conjunction with standard methods in order to maximize obtainable information for better management of patients with MR.

Chromosomal alterations in osteosarcoma cell lines revealed by comparative genomic hybridization and multicolor karyotyping

We characterized the chromosomal alterations in eight osteosarcoma cell lines (OST, HOS, U-2 OS, ZK-58, MG-63, SJSA-1, Saos-2, and MNNG) by comparative genomic hybridization (CGH); gains and losses of DNA sequences were defined as chromosomal regions with a fluorescence ratio, wherein all of the 95% confidence interval was above 1.25 and below 0.75, respectively. In four of 8 cell lines, multicolor karyotyping (MK) was added. CGH revealed the average number of aberrations per cell line was 20.8 (range: 10–31); the average numbers of gains and losses were 11.1 and 9.6, respectively. The frequent gains were identified on 1p21∼q24, 1q25∼q31, 7p21, 7q31, 8q23∼q24, and 14q21; frequent losses were at 18q21∼q22, 18q12, 19p, and 3p12∼p14. High-level gains were observed on 8q23∼q24, 5p, and 1p21∼p22. MK revealed the most common translocations in the four cell lines were t(8;9), t(1;3), t(3;5), t(1;13), t(2;6), t(3;17), t(1;15), t(10;20), and t(6;20). Chromosomes 1, 3, 8, 9, and 20 were most frequently involved in translocation events. The concordance rate of aberrations in CGH and translocations in MK was 76%. MK was useful to identify the chromosomal alterations and as a supplement to the CGH results in three of four chromosomes.

Multicolor Karyotyping in Acute Myeloid Leukemia

Cytogenetic data have significantly contributed to our understanding of the heterogeneity of acute myeloid leukemia (AML). In AML, numerous recurrent chromosomal aberrations have been identified, and several of them, e.g. t(8;21)(q22;q22), t(15;17)(q22;q11-12), inv(16)(p13q22), are specific for distinct subgroups. Furthermore, chromosomal aberrations have proved to be of paramount prognostic importance for remission induction and survival. Chromosome analysis using classical cytogenetic banding techniques often fails to completely resolve complex karyotypes and cryptic translocations not identifiable by these techniques have been detected using molecular cytogenetic methods. While fluorescence in situ hybridization (FISH) has become an indispensable tool for screening and follow-up of known aberrations, the techniques of spectral karyotyping (SKY) and multiplex-fluorescence in situ hybridization (M-FISH) allow for the simultaneous visualization of all chromosomes of a metaphase in a single hybridization step, and thereby enable screening for the aberrations present without their prior knowledge. Therefore, with the introduction of these techniques in 1996 the comprehensive analysis of complex karyotypes and the identification of new, hitherto cryptic translocations and, ultimately, the identification of new disease subgroups seemed possible. Since, more than 600 cases of AML and MDS have been analyzed. Herein, we attempt to summarize the data published and discuss what has been achieved towards realization of these goals.