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• W2402818607 abstract "This issue features nine original manuscripts in the field of clinical cytometry covering a fairly wide range of topics, which may be summarized as follows: Three papers are from the field of leukemia/lymphoma immunophenotyping. First, Preijers and colleagues 1 review the results and lessons learned from 15 years of external quality assurance (EQA) in The Netherlands and Belgium. They distributed 90 samples dominated by aberrant cells with the requests to characterize them and formulate a brief, clinically oriented conclusion. In order to analyze the data, the outcomes of the pre-analytical, analytical and post-analytical assay phases were scored as correct or having minor or major faults. This approach was inspired by the standardization efforts of the International Council for Standardization in Hematology (ICSH) and the International Clinical Cytometry Society (ICCS) 2-4. These condensated results allowed some appreciation of trends over time pointing towards the continuing need to improve technical aspects of the assay, and improved interpretational skills of the participants. An important logistic trend was observed, namely that smaller peripheral centers (i.e., serving hospitals with <600 beds) did less well than the larger peripheral centers, whilst the academic centers did best. Although retrospective, this study is a strong plea to concentrate leukemia/lymphoma immunophenotyping in specialized centers. Cavazzini et al. 5 addressed CD146 expression by acute leukemia blast cells in a single-center study. CD146, known as melanoma cell adhesion molecule (MCAM), is rarely expressed on normal human hematopoietic cells. Here, CD146 was rarely expressed by acute myelogenous leukemia (AML, n=121), but more frequent by B-acute lymphoblastic leukemia (ALL, n=38) and by 2 of 3 T-ALL cases. In addition to Gaipa's review of minimal residual disease (MRD) in ALL 6, Cavazzini's pilot study suggest that in some cases of ALL, CD146 might be useful as a marker for the follow-up of this disease. [Color figure can be viewed in the online issue, which is available at wileyonlinelibrary.com.] Epithelial cells can be distinguished from leukocytes on the basis of their immunophenotype (i.e., CD45neg, CD326+ vs. CD45+, CD326neg, respectively). In body fluids (such as pleural, peritoneal or pericardial), the additional use of CD33 allows further distinction from other cell types such as monocytes or macrophages (CD45+, CD33+, CD326neg) and mesothelial cells (CD45dim, CD33neg, CD326neg). Acosta and colleagues 9 used this panel to characterize 238 body fluid samples in comparison to “classical” approaches, i.e., morphology on smears and cell blocks, and immunochemistry paraffin wax embedded blocks. Flow cytometry had very good sensitivity and specificity (>95%). This study extended the work of Kitiyama et al. 10 on ascites and peritoneal lavages where tumor cells and leukocytes were distinguished in a similar way. Two papers from the Locus Medicus group in Athens addressed the contribution that flow cytometry can make to detect M. tuberculosis infection (MTB) 11 or to quantify immune responses against this bacterium 12. Early secreted antigenic target (ESAT)-6 is a marker of virulence for MTB. The Locus Medicus group developed an assay to detect ESAT-6 using intracellular flow cytometry and demonstrated the feasibility of this assay 11. Immune responses against MTB were quantified flow cytometrically using induction of intracellular IFNγ in CD4+ T cells after stimulation with ESAT-6 or tuberculin purified protein derivate (PPD) 12. An enzyme-linked immunosorbent assay (ELISA; Quantiferon TB Gold In-Tube) and the tuberculin skin test served as references. Five groups were studied; patients with active MTB, close contacts to active MTB patients, patients with rheumatic disease under treatment with biologic agents, age-matched control persons for the latter group, and healthy controls naïve to MTB. The frequencies of positive test results were clearly higher in CD4+IFNγ+ cells after stimulation with ESAT-6 or PPD than in ELISA or after skin testing. Similar proportions of positive test results were observed in the three patient groups for the 2 CD4+IFNγ+-based assays. Earlier, Kim and colleagues 13 had reported that flow cytometry produced more diverse information on dynamic changes in the CD4+IFNγ+ T-cell compartment in patients with active MTB receiving anti-tuberculous therapy. The diagnosis of sepsis by blood culture typically requires 2-3 days which is not fast enough in neonates in whom bacterial infection may spread rapidly with a high risk for mortality. Earlier, Park et al. 14 showed, in adult patients, that a 16-part differential count correlated with sepsis severity (i.e., significant reductions in absolute counts of all lymphocyte subsets in patients with complicated sepsis vs. those with uncomplicated sepsis). In neonatal patients, Pradhan et al. 15 demonstrated the usefulness of flow cytometric quantification of elevated CD64 intensity on neutrophils to diagnose sepsis, whilst in the same cohort reduced intensity of HLA-DR on monocytes represented a simultaneously occurring anti-inflammatory response. The combination of the intensity of expression of both parameters in the form of a “sepsis index” added some diagnostic sensitivity to neutrophil CD64 intensity to detect neonatal sepsis. The diagnosis of allergic reactions is based on clinical history and the demonstration of specific IgE. Basophils may release histamine following allergen-mediated crosslinking of FcεRI-specific IgE on these cells. Classical basophil histamine release tests have nowadays been replaced by flow cytometric basophil activation testing 16. These tests are based on the upregulation of expression of CD63 (barely detectable on resting basophils) and CD203c (constitutively expressed). Histamine and its release can also be studied by an enzyme affinity method based on the affinity of the histaminase diamine oxidase (DAO) for its substrate histamine (termed “HistaFlow”) 17. Here, Cop et al. 18 extended the applications of HistaFlow from birch pollen allergy 17 and wasp venom immunotherapy 19 to that of a small chemical that generally elicits little basophil activation, i.e., rocuronium allergy. Thus, histamine release by individual cells in response to drugs can be studied by flow cytometry upon IgE-dependent and IgE-independent challenges. The diagnosis of hemoglobin disorders is typically made on the detection of abnormal red blood cell parameters, pathological hemoglobin types by high-pressure liquid chromatography and confirmation by molecular biology. Whilst screening of large amounts of samples is routinely done by the naked eye using the single-tube red cell osmotic fragility test, Mohapatra and colleagues, extending their previous work 20 propose a flow cytometric alternative (21). Here, EDTA-anticoagulated blood is first diluted in saline, after which forward light scatter is acquired against time. After initial acquisition, the sample is spiked with deionized distilled water, and the proportion of residual red cells quantified and compared with that of the unspiked list mode data. The naked eye had a similarly high sensitivity and specificity to detect both α-and β-thalassemia traits, but missed the majority of HbS carriers. Flow cytometry was most sensitive to discriminate hemoglobinopathy carriers from normals, irrespective of the subtype." @default.
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• W2402818607 date "2016-05-01" @default.
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• W2402818607 title "Issue highlights - May 2016" @default.
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• W2402818607 doi "https://doi.org/10.1002/cyto.b.21379" @default.
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