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Conventional G-banding cytogenetics (CC) detects chromosome 17 (chr17) abnormalities in 2% of patients with de novo myelodysplastic syndromes (MDS). We used CC and fluorescence in situ hybridization (FISH) (LSI p53/17p13.1) to assess deletion of 17p in 531 patients with de novo MDS from the Spanish Group of Hematological Cytogenetics. FISH detected - 17 or 17p abnormalities in 13 cases (2.6%) in whom no 17p abnormalities were revealed by CC: 0.9% of patients with a normal karyotype, 0% in non-informative cytogenetics, 50% of patients with a chr17 abnormality without loss of 17p and 4.7% of cases with an abnormal karyotype not involving chr17. Our results suggest that applying FISH of 17p13 to identify the number of copies of the TP53 gene could be beneficial in patients with a complex karyotype. We recommend using FISH of 17p13 in young patients with a normal karyotype or non-informative cytogenetics, and always in isolated del(17p).

Chromosomal abnormalities are detected in 40-60% of patients with de novo myelodysplastic syndromes (MDS). This study used the FISH technique in 773 patients with de novo MDS without evidence of monosomy 7 (-7) or 7q deletion (7q-) by conventional G-banding cytogenetics (CC) to analyze their prognostic impact by FISH alone. FISH detected -7/7q- in 5.2% of patients. Presence of -7/7q- was associated with shorter overall survival than absence of such aberrations. Our results suggest that FISH 7q could be beneficial in patients with intermediate WHO morphologic risk stratification and no evidence of -7/7q- by CC., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Background and Objectives: Despite the great utility of chimerism analysis after allogeneic stem cell transplantation, a gold standard method for its quantification has not yet been defined. The objective of the present investigation was to compare the sensitivity (detection limit) and the quantification accuracy of fluorescent in situ hybridization with specific probes for the sex chromosomes (XY-FISH) and multiplex short tandem repeat polymerase chain reaction (STR-PCR) revealed by capillary electrophoresis for the quantification of chimerism after stem cell transplantation., Design and Methods: A first experiment was performed on two sets of artificial cell mixtures from two sex-mismatched healthy donors mixed in different proportions (% male: 100, 75, 50, 25, 10, 5, 3, 1, 0.1, 0). In a second experiment, 58 samples obtained from 10 selected patients with different clinical courses and chimerism evolution after sex-mismatched stem cell transplantation, which had been studied by XY-FISH, were retrospectively analyzed by STR-PCR. In a third experiment, 60 unselected prospective samples belonging to 15 patients (5 of whom had also been included in the retrospective study) were analyzed by both XY-FISH and STR-PCR., Results: Both techniques showed high quantification accuracy and were highly reproducible. The sensitivity of both approaches reached 1% under standard conditions. Moreover, the use of long injection times for the capillary electrophoresis (30 and 50s vs. the standard 10s) resulted in an increase of sensitivity of the STR-PCR assay up to 0.1%, which has interesting clinical implications., Interpretation and Conclusions: Considering the high sensitivity and quantification accuracy of multiplex STR-PCR and the fact that this assay is sex-independent and can be applied to virtually all patients, STR-PCR could be considered as the method of choice for chimerism quantification after stem cell transplantation when high sensitivity is not a requirement.

Background: Fluorescence in situ hybridisation (FISH) is useful for detecting specific chromosomal abnormalities in various tumours. In lymphomas, diagnosis is frequently made using paraffin wax embedded tissue. However, FISH performed under these conditions presents potential technical problems and difficulties in interpretation., Aims: To show that FISH using tissue imprints and cytopreps or alternatively, bone marrow (BM) smears, constitutes an easy and rapid strategy to overcome these constraints., Methods: The study comprised 46 patients with lymphoma. Sixty nine tissue imprints, cytopreps, or BM smears were analysed by FISH. Dual colour, dual fusion FISH probes were used to detect the t(8;14), t(11;14), and t(14;18) translocations, whereas a dual colour breakapart FISH probe was used to detect chromosomal translocations involving the BCL6 gene., Results: Tissue imprints and cytopreps were successfully hybridised in all 52 cases, whereas hybridisation was successful in 16 of 17 archival BM smears. All patients could be analysed to identify either the presence or absence of chromosomal translocations., Conclusions: The use of tissue imprints, cytopreps, or BM smears to identify chromosomal abnormalities by FISH is a rapid and useful ancillary approach for diagnostic purposes. Therefore, it could be used on a routine basis whenever fresh samples are available.

The usefulness of fluorescence in situ hybridization (FISH) analysis to detect minimal residual disease (MRD) in autologous bone marrow and peripheral blood stem-cell harvests has been tested in three patients with hematologic malignancies. Conventional cytogenetics and FISH were used to characterize the leukemic clones identifying the specific chromosomal abnormalities (monosomy 7 in a myelodysplastic patient and trisomy 8 in two acute myeloid leukemic patients). Such analysis was useful to monitor the MRD persistent after treating these patients with intensive chemotherapy. The myelodysplastic patient underwent eight peripheral blood-stem cell harvests in which FISH detected the persistence of monosomy 7 cells, precluding their use for autologous transplantation. This patient relapsed and died. In two acute myeloid leukemia patients who underwent an autologous marrow harvest, FISH did not show a significant proportion of trisomy 8 cells. Nevertheless, autologous transplantation was not performed, owing to an insufficient CD34 cell content in the harvests. One of these patients relapsed with the reappearance of trisomy 8 and died. The other patient, on the contrary, is alive in complete remission 3 years after the bone marrow harvest. The usefulness and applicability of MRD quantification in stem-cell harvests is discussed on the basis of the sensitivity of the methodology applied.

Using a highly sensitive fluorescence in situ hybridization method with probes for BCR and ABL1 (D-FISH), we studied 37 paired sets of bone marrow and blood specimens, collected within 24 to 96 hours of each other, from 10 patients before and during treatment for chronic myeloid leukemia (CML). The normal range for 500 interphase nuclei was

The efficiency of restriction endonuclease in situ digestion (REISD) with Sau3A to analyze chimerism and residual disease (RD) has been tested before and after an allogenic bone marrow transplant (BMT) in an acute lymphoblastic leukemia (ALL) patient. The combined results obtained with REISD and FISH using the appropriate probes for detecting chromosome rearrangements have proven to be useful for the identification and quantification of both the hemopoietic chimerism achieved after BMT and the RD persistent in the patient. The sensitivity of REISD has been determined to be around 95%, i.e., similar to that obtained by FISH. REISD with Sau3A was particularly useful in the analysis of chimerism since this enzyme revealed the polymorphic status of constitutive heterochromatin in human chromosome 3 and thus allowed discrimination of cells derived from donor and recipient. The method itself seems promising since neither a donor/recipient sex mismatch nor a cytogenetic disease marker are needed for its application.

Fluorescence in situ hybridization (FISH) allows detection of the intercellular heterogeneity of C-ERB-B2 gene amplification in uncultured breast cancer cells. Nevertheless, because high levels of amplification result in coalescence of signals, direct microscopy quantification is restricted to cells wih low levels of amplification or with dispersed signals. A methodology of digital image analysis, using surface and grey-level FISH signals as parameters that permit a rapid, objective, and accurate estimation of gene copy number, is presented. This procedure is independent of the signal overlapping and results in a more accurate quantification and characterization of tumor cell heterogeneity.