Automated four-color interphase fluorescence in situ hybridization approach for the simultaneous detection of specific aneuploidies of diagnostic and prognostic significance in high hyperdiploid acute lymphoblastic leukemia

High hyperdiploid acute lymphoblastic leukemia in adults shows clonal heterogeneity and chromosomal instability at diagnosis and during the course of the disease

“…Automated four-color interphase fluorescence in situ hybridization (I-FISH) previously revealed a high level of clonal aneuploidy heterogeneity in HeH ALL at presentation [4]. Numerical chromosome instability (CIN) was supposed to be at the origin of this heterogeneity.…”

“…Average CIN was determined for all four chromosomes together and then for each selected chromosome. Approaches used and cutoff levels were reported in detail previously [4,7]. Table 1 Clones involving chromosomes 4, 6, 10, and 17 at presentation and/or during the course of disease in ten patients with high hyperdiploid acute lymphoblastic leukemia, expressed as percentages (13) 29 (13) 24 (15) 13 (8) 18 (15) 44 (34) 6 (3) 8 (5) 19 (16) 40 (25) 7 (7) 3 (3) 20 (13) 56 (43) 12 (9) 2 (1) 8 (5) 7 (6) Values in parentheses are numbers of very small abnormal clones (<1%).…”

“…Study patients are further detailed in Supplement 1. Four-color I-FISH using centromeric probes specific for chromosomes 4 (p-4n1/4, kindly provided by Prof. Mariano Rocchi, University of Bari, Italy), 6, 10, and 17 (D6Z1, D10Z1, and D17Z1, respectively; American Type Culture Collection, Manassas, VA) was performed as formerly described [4] on 34 bone marrow samples, 5 of which were reported in Talamo et al [7]. Probes were directly labeled by nick translation with four different fluorochromes (Cy3, Cy3.5, DEAC, and FITC).…”

State‐of‐the‐art FISHing: Automated analysis of cytogenetic aberrations in interphase nuclei

“…As an example, Blandin et al investigated aneusomy of chromosomes 4, 6, 10, and 17 in high hyperdiploid acute lymphoblastic leukemia using four-color FISH. The large number of nuclei scored revealed a high level of chromosome variability both at diagnosis and relapse, which might have prognostic significance and huge effect on future treatment strategies (42).…”

“…To detect FISH spots, a 320 objective might be appropriate (28,29); however, if not only the number and intensity of signals are important but their spatial pattern as well, the application of a higher magnification and more importantly high resolution is desirable (30)(31)(32). Considering the prior, several workgroups use 360, 363, or 3100 objectives during the automated i-FISH analysis (21,(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43). An alternative approach, also increasing speed, is the use of two objectives, one of them for nucleus selection (objective magnification: and the other one for detecting FISH signals (objective magnification: 340-100) (31,44-52).…”