SummaryAcute leukaemias of ambiguous lineage (ALAL) represent a rare type of leukaemia, expressing both myeloid and lymphoid markers. This study retrospectively analyzed data from 92 children (biphenotypic n = 78, bilineal n = 6, lineage switch n = 8) with ALAL registered in the Berlin-Frankfürt-Münster (BFM) acute myeloid leukaemia (AML) and acute lymphoblastic leukaemia (ALL) studies between 1998 and 2006 (2AE4% of all cases with acute leukaemia). Our cohort of ALAL patients was characterized by comparatively high median age (8AE9 years), high median white blood cell count (14AE9 · 10 9 /l), as well as frequent hyperleucocytosis (18AE5%) and central nervous system involvement (24AE1%). The most frequent cytogenetic abnormalities were ETV6/RUNX1 fusion (16%) and trisomy 8 (14AE6%). Complete remission rate was significantly lower than in ALL-BFM patients (91AE8% vs. 99AE1%, P < 0AE001), but comparable to AML-BFM patients (87AE9%). Event-free survival (EFS) and overall survival (OS) of ALAL patients were low, at 62 ± 5%. 5-year probability of EFS was significantly worse than in ALL patients (80 ± 1%, P < 0AE001), but better than for AML patients (49 ± 2%, P = 0AE027). Our data suggest that an intensive therapy regimen including stem cell transplantation may be favourable for bilineal or lineage switch cases, whereas patients with ETV6/RUNX1 fusion, lymphoid morphology and patients with expression of cyCD22 and cyCD79a should be treated with an ALL-directed therapy.

Fluorescence in situ hybridisation (FISH) has gained major impact in the detection of chromosomal aberrations. The application of FISH, however, is hampered due to complicated protocols. We therefore designed a FISH protocol that allows the fast and reliable application on cytological and histological specimens. Cytological and histological specimens of bone marrow, lymph nodes, liver and breast were analysed with 14 sets of centromere or locus-specific probes. A pretreatment of enzymatic digestion and microwave heating with the same incubation times for all specimens and probes used was performed. Hybridisation efficiency was proved by calculating the thresholds for monosomy, trisomy and translocations. Values in cytological specimens reached 9.0, 3.9 and 4.6%, respectively. In histological sections, values of 6.4% were seen for aneusomy and 3.1% for translocations. However, values for monosomy reached 20.5% in bone marrow and 61.8% in liver tissues due to cutting artifacts. In bone marrow with acute myeloid leukaemias, lymph nodes with follicular and mantle cell lymphomas, breast carcinomas and liver cell carcinomas, FISH confirmed aberrations already found using conventional cytogenetics. The protocol shown here is easy to perform and can be used with cytological and histological specimens. Moreover, with "hands on" time of less than 2 h, FISH can also be used for daily routine purposes.

In this study children experienced less pain after 60 min application time, but pain reduction was already seen after 40 min. The child's perception of pain differed from observers' point of view and should therefore always be included in pain management.

Objectives: Fluorescence in situ hybridization (FISH) has become a useful tool to identify chromosomal aberrations in non-dividing cells. Numerous studies have compared chromosomal banding analysis (CBA) and FISH on fixed cultured bone marrow cells. However, up to now, there has been no study comparing two main sources of diagnostic material, i.e. bone marrow aspirates and trephine biopsies. We therefore analyzed these materials by FISH in comparison with CBA. Methods: CBA revealed chromosomal aberrations in 18 patients suffering from myelodysplastic syndrome (n = 13), acute myeloid leukemia (n = 3), or chronic myeloproliferative syndrome (n = 2). FISH was performed on fixed cultured bone marrow cells, aspirates and trephine biopsies from each patient. Results: Percentages of aberrant cells in the different materials correlated highly with Pearson values of 0.909 for biopsy/fixed cultured cells (p < 0.001), 0.830 for biopsy/aspirate (p < 0.001) and 0.768 for aspirate/fixed cultured cells (p < 0.001). Moreover, in bone marrow biopsies peritrabecular and central intertrabecular areas yielded very similar FISH results with a high correlation (r = 0.968, p < 0.001). FISH revealed a lower proportion of aberrant cells than CBA in 90% of the specimens. Conclusions: In summary, the different materials available for the FISH examination are comparable in sensitivity and show similar quantitative results. Therefore, the use of biopsy sections for the routine FISH examination of chromosomal abnormalities is a valid method.

The PRDX4 gene located at Xp22 encodes for a member of the peroxiredoxin gene family. Genes within this family exhibit thioredoxin-dependent peroxidase activity and have been implicated in cellular functioning, including proliferation and differentiation. Recently, PRDX4 has been identified as a partner gene in a t(X;21) translocation in a patient with acute myeloid leukemia. To determine whether PRDX4 was involved in other translocations, leukemia cells from 15 patients with Xp22 abnormalities were screened for involvement of the gene using fluorescence in situ hybridization (FISH). One sample from a 41-year-old woman with acute lymphoblastic leukemia showed three signals when hybridized with the PRDX4 probe. Cytogenetic analysis of the sample had identified a t(X;18)(p22;q23). Assuming that the three signals indicated a break within the PRDX4 gene, we performed FISH experiments and successfully narrowed the breakpoint on chromosome 18 to a 50-kb region. Subsequent analysis using spectral karyotyping showed that the leukemic cells had undergone multiple rearrangements and that a third X chromosome was present, albeit rearranged. Additional FISH experiments revealed that the third PRDX4 signal was the result of a third copy of the gene. Analysis of the other rearrangements has helped to characterize the multiple abnormalities within the leukemic cells. The findings underscore the importance of using multiple techniques when analyzing complex chromosomal rearrangements in malignant cells.