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Introduction: Flow cytometric panels for the investigation of lymphoproliferative disorders, such as the EuroFlow Lymphoid Screening Tube (LST), often fail to demonstrate T‐cell clonality, as a suitable clonality marker was unavailable until recently. Aim of this study was to evaluate the added value of supplementing TRBC1, a flow cytometric T‐cell clonality marker, to the LST. Methods: Flow cytometric analysis was performed on 830 routine samples referred to our lab for suspicion of hematological malignancy. T‐cells with monotypic TRBC1‐expression were additionally characterized with a 12‐color T‐cell tube and molecular T‐cell receptor gamma gene rearrangement (TRG). Results: LST analysis revealed 97 (11.7%) samples with the presence of a monotypic T‐cell population according to TRBC1, including 21 (2.5%) "high‐count" (≥500 cells/μL blood or ≥15% of lymphocytes) and 76 (9.2%) "low‐count" (<500 cells/μL blood or <15% of lymphocytes) populations. Clinical symptoms indicative for T‐CLPD could be correlated to 11/21 "high‐count" and 17/76 "low‐count" monotypic T‐cell populations. Molecular TRG analysis demonstrated a monoclonal result in 76% (16/21) of "high‐count" samples and in 64% (42/66; 10 samples not tested) of "low‐count" samples, but also in 9/20 samples with polytypic TRBC1 results. Conclusion: Analysis of an LST tube supplemented with TRBC1 led to the detection of a high number of monotypic T‐cell populations. The detection of numerous small monotypic T‐cell populations raises the question of their clinical significance. A possible flowchart for assessment of these populations, based on the available literature, is proposed. Molecular TRG analysis is complementary and cannot be omitted from T‐cell clonality assessment. [ABSTRACT FROM AUTHOR]

Introduction Routine hematology parameters in hematopoietic progenitor cell apheresis products ( HPC-A) are usually determined using automated cell counters. These instruments, however, are designed to analyze whole blood samples, that differ considerably from HPC-A in blood cell composition. This study evaluates the performance of two automated cell counters for the analysis of HPC-A. Methods Routine hematology parameters [red blood cells ( RBC), hematocrit ( HCT), mean corpuscular volume ( MCV), white blood cells ( WBC), WBC differentiation, and platelets ( PLT)] were determined on the Unicel DxH 800 instrument (Beckman Coulter) and the XN-350 instrument (Sysmex). Correlations with the reference methods, intrarun precision, and linearity of the analyses were studied. Results Good correlations were found for almost all parameters. However, RBC count was overestimated by XN-350, using the impedance technique, as was neutrophil percentage using DxH 800. Coefficients of variation for intrarun precision were below 10% on both analyzers for all parameters, except for neutrophil percentage (14.7%) and PLT (10%) on DxH 800. Both instruments showed good linearity for all parameters, except for RBC and HCT on DxH 800. Conclusion With the exception of the measurement of neutrophils on DxH 800 and RBC by the impedance technique on the XN-350, routine hematology parameters in HPC-A can safely be determined using automated cell counters. [ABSTRACT FROM AUTHOR]

Introduction Standardization of BCR- ABL1 messenger RNA quantification by real-time PCR on the International Scale ( IS) is critical for monitoring therapy response in chronic myelogenous leukaemia. Since 2006, BCR- ABL1 IS standardization is propagated along reference laboratories by calculating a laboratory-specific conversion factor ( CF), co-ordinated in Europe through the European Treatment and Outcome Study project. Although this process has proven successful to some extent, it has not been achievable for all laboratories due to the complexity of the process and the stringent requirements in terms of numbers of samples to be exchanged. In addition, several BCR- ABL1 IS quantification methods and secondary reference materials became commercially available. However, it was observed that different IS methods generate consistently different results. Methods To overcome these difficulties, we have developed an alternative and simple approach of CF calculation, based on the retrospective analysis of existing external quality assessment ( EQA) data. Our approach does not depend on the exchange of samples and is solely based on the mathematical CF calculation using EQA results. Results and conclusion We have demonstrated by thorough statistical validation that this approach performs well in converting BCR- ABL1 measurements to improve IS estimation. In expectation of a true golden standard method for BCR- ABL1 IS quantification, the proposed method is a valuable alternative. [ABSTRACT FROM AUTHOR]

Introduction The CellaVision Advanced Red Blood Cell ( RBC) Software Application is a new software for advanced morphological analysis of RBCs on a digital microscopy system. Upon automated precharacterization into 21 categories, the software offers the possibility of reclassification of RBCs by the operator. We aimed to define the optimal cut-off to detect morphological RBC abnormalities and to evaluate the precharacterization performance of this software. Methods Thirty-eight blood samples of healthy donors and sixty-eight samples of hospitalized patients were analyzed. Different methodologies to define a cut-off between negativity and positivity were used. Sensitivity and specificity were calculated according to these different cut-offs using the manual microscopic method as the gold standard. Imprecision was assessed by measuring analytical within-run and between-run variability and by measuring between-observer variability. Results By optimizing the cut-off between negativity and positivity, sensitivities exceeded 80% for 'critical' RBC categories (target cells, tear drop cells, spherocytes, sickle cells, and parasites), while specificities exceeded 80% for the other RBC morphological categories. Results of within-run, between-run, and between-observer variabilities were all clinically acceptable. Conclusion The CellaVision Advanced RBC Software Application is an easy-to-use software that helps to detect most RBC morphological abnormalities in a sensitive and specific way without increasing work load, provided the proper cut-offs are chosen. However, evaluation of the images by an experienced observer remains necessary. [ABSTRACT FROM AUTHOR]

Introduction The CellaVision Advanced Red Blood Cell ( RBC) Software Application is a new software for advanced morphological analysis of RBC, which automatically performs a preliminary characterization and grouping of RBC into 21 morphological categories, including schistocytes. Upon automated classification, the software offers the possibility of reclassification of RBC by the operator. The aim of this study was to evaluate the schistocyte analysis by the CellaVision Advanced RBC Application. Methods Schistocyte counts were evaluated comparing the automated count on a CellaVision DM96, both before and after reclassification, with the reference manual microscopic method according to the ICSH criteria. Thirty-six samples of hospitalized patients and 40 samples of controls were analyzed. Results Within-run, between-run and between-observer coefficients of variation were lower when counted with the CellaVision compared to the manual microscopic count. The very high sensitivity but rather poor specificity implicates the need for reclassification by the operator, following automated analysis. After reclassification, method comparison studies revealed good agreement with the manual microscopic method, with however slightly higher values of schistocytes for the automated analysis. Conclusion The CellaVision Advanced RBC Software Application provides a sensitive and reproducible measurement of schistocytes in peripheral blood, but still requires manual revision. Furthermore, it is an easy-to-use software and an excellent teaching tool that might contribute to standardization in the investigation of schistocyte-related conditions. [ABSTRACT FROM AUTHOR]

Recently, we and others described a new chromosomal rearrangement, that is, inv(7)(p15q34) and t(7;7)(p15;q34) involving the T-cell receptor beta (TCRβ) (7q34) and the HOXA gene locus (7p15) in 5% of T-cell acute lymphoblastic leukemia (T-ALL) patients leading to transcriptional activation of especially HOXA10. To further address the clinical, immunophenotypical and molecular genetic findings of this chromosomal aberration, we studied 330 additional T-ALLs. This revealed TCRβ-HOXA rearrangements in five additional patients, which brings the total to 14 cases in 424 patients (3.3%). Real-time quantitative PCR analysis for HOXA10 gene expression was performed in 170 T-ALL patients and detected HOXA10 overexpression in 25.2% of cases including all the cases with a TCRβ-HOXA rearrangement (8.2%). In contrast, expression of the short HOXA10 transcript, HOXA10b, was almost exclusively found in the TCRβ-HOXA rearranged cases, suggesting a specific role for the HOXA10b short transcript in TCRβ-HOXA-mediated oncogenesis. Other molecular and/or cytogenetic aberrations frequently found in subtypes of T-ALL (SIL-TAL1, CALM-AF10, HOX11, HOX11L2) were not detected in the TCRβ-HOXA rearranged cases except for deletion 9p21 and NOTCH1 activating mutations, which were present in 64 and 67%, respectively. In conclusion, this study defines TCRβ-HOXA rearranged T-ALLs as a distinct cytogenetic subgroup by clinical, immunophenotypical and molecular genetic characteristics.Leukemia (2007) 21, 121–128. doi:10.1038/sj.leu.2404410; published online 12 October 2006 [ABSTRACT FROM AUTHOR]

Chromosomal aberrations of T-cell receptor (TCR) gene loci often involve the TCRαδ (14q11) locus and affect various known T-cell oncogenes. A systematic fluorescent in situ hybridization (FISH) screening for the detection of chromosomal aberrations involving the TCR loci, TCRαδ (14q11), TCRβ (7q34) and TCRγ (7p14), has not been conducted so far. Therefore, we initiated a screening of 126 T-cell acute lymphoblastic leukemia (T-ALL) and T-cell lymphoblastic lymphoma cases and 19 T-ALL cell lines using FISH break-apart assays for the different TCR loci. Genomic rearrangements of the TCRβ locus were detected in 24/126 cases (19%), most of which (58.3%) were not detected upon banding analysis. Breakpoints in the TCRαδ locus were detected in 22/126 cases (17.4%), whereas standard cytogenetics only detected 14 of these 22 cases. Cryptic TCRαδ/TCRβ chromosome aberrations were thus observed in 22 of 126 cases (17.4%). Some of these chromosome aberrations target new putative T-cell oncogenes at chromosome 11q24, 20p12 and 6q22. Five patients and one cell line carried chromosomal rearrangements affecting both TCRβ and TCRαδ loci. In conclusion, this study presents the first inventory of chromosomal rearrangements of TCR loci in T-ALL, revealing an unexpected high number of cryptic chromosomal rearrangements of the TCRβ locus and further broadening the spectrum of genes putatively implicated in T-cell oncogenesis.Leukemia (2006) 20, 1238–1244. doi:10.1038/sj.leu.2404243; published online 4 May 2006 [ABSTRACT FROM AUTHOR]

A letter to the editor is presented in response to the article "HOXA Cluster Deregulation in T-ALL Associated With Both a TCRD-HOXA and a CALM-AF10 Chromosomal Translocation," that was published in the March 2006 issue.

Chromosomal translocations with breakpoints in T-cell receptor (TCR) genes are recurrent in T-cell malignancies. These translocations involve the TCRadgene (14q11), the TCRßgene (7q34) and to a lesser extent the TCR?gene at chromosomal band 7p14 and juxtapose T-cell oncogenes next to TCR regulatory sequences leading to deregulated expression of those oncogenes. Here, we describe a new recurrent chromosomal inversion of chromosome 7, inv(7)(p15q34), in a subset of patients with T-cell acute lymphoblastic leukemia characterized by CD2 negative and CD4 positive, CD8 negative blasts. This rearrangement juxtaposes the distal part of the HOXA gene cluster on 7p15 to the TCRßlocus on 7q34. Real time quantitative PCR analysis for all HOXA genes revealed high levels of HOXA10 and HOXA11 expression in all inv(7) positive cases. This is the first report of a recurrent chromosome rearrangement targeting the HOXA gene cluster in T-cell malignancies resulting in deregulated HOXA gene expression (particularly HOXA10 and HOXA11) and is in keeping with a previous report suggesting HOXA deregulation in MLL-rearranged T- and B cell lymphoblastic leukemia as the key factor in leukaemic transformation. Finally, our observation also supports the previous suggested role of HOXA10 and HOXA11 in normal thymocyte development.Leukemia (2005) 19, 358-366. doi:10.1038/sj.leu.2403657 Published online 27 January 2005 [ABSTRACT FROM AUTHOR]

The emergence of heterogeneous populations of methicillin-resistant Staphylococcus aureus (MRSA) causes major problems in routine screening for MRSA. In heterogeneous MRSA populations, a proportion of bacterial cells show low-level resistance to oxacillin, with minimal inhibitory concentrations (MICs) of oxacillin ranging between 1 and 100 mg/l, while in homogeneous MRSA populations, the MIC of oxacillin for all cells is >100 mg/l. Routine oxacillin disk diffusion tests often fail to detect heterogeneous MRSA populations. In the present study, a recently proposed disk diffusion method that employs a cephamycin antibiotic (cefoxitin 30 µg; BD Sensi-disc, Becton Dickinson, Germany) was evaluated using 155 clinical isolates of S. aureus (73 mecA positive and 82 mecA negative). The results were compared with those of other MRSA screening techniques: a disk diffusion test with oxacillin 1 µg and cefoxitin 30 µg (BD Sensi-disc; Becton Dickinson), an MRSA latex agglutination tests (Denka Seiken, Japan), and an oxacillin screen agar test (6 µg/ml; Becton Dickinson). Detection of the mecA gene by polymerase chain reaction was considered the gold standard. The performances of the different methods were determined and compared. The results showed that the cefoxitin disk diffusion test is preferable to the oxacillin disk diffusion method for routine screening to detect MRSA. [ABSTRACT FROM AUTHOR]

Examines the condition of bone marrow and peripheral blood in patients with myelodysplastic syndrome with monoclonal B lymphocytes. Inactivation of X-chromosomes; Origin of malignant cell lines from transformed pluripotent stem cell; Detection of hypercellular smear with hyperplastic dyserythropoiesis in bone marrow.

HLA-C*05:99N results from a single nucleotide loss compared with its closest allele HLA-C*05:01:01:01. [ABSTRACT FROM AUTHOR]

Background: Variant translocations t(9;22) occur in 5 to 10% of newly diagnosed CMLs and additional genetic changes are present in 60-80% of patients in blast crisis (BC). Here, we report on a CML patient in blast crisis presenting with a four-way variant t(9;22) rearrangement involving the EVI1 locus.Methods: Dual-colour Fluorescence In Situ Hybridisation was performed to unravel the different cytogenetic aberrations. Expression levels of EVI1 and BCR/ABL1 were investigated using real-time quantitative RT-PCR.Results: In this paper we identified a patient with a complex 4-way t(3;9;17;22) which, in addition to BCR/ABL1 gene fusion, also resulted in EVI1 rearrangement and overexpression.Conclusion: This report illustrates how a variant t(9;22) translocation can specifically target a second oncogene most likely contributing to the more aggressive phenotype of the disease. Molecular analysis of such variants is thus warranted to understand the phenotypic consequences and to open the way for combined molecular therapies in order to tackle the secondary oncogenic effect which is unresponsive to imatinib treatment. [ABSTRACT FROM AUTHOR]