Flow Cytometric Analysis of Acute Leukemias
Zahid KaleemEric CrawfordM. Hanif PathanLeah JasperMichael A. CovinskyLawrence R. JohnsonG. Wesley White
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Abstract Context.—Acute leukemia displays characteristic patterns of surface antigen expression (CD antigens), which facilitate their identification and proper classification and hence play an important role in instituting proper treatment plans. In addition to enzyme cytochemical analysis, multiparameter flow cytometric analysis has become commonplace in most laboratories for that purpose. The essential role and caveats of flow cytometry in that regard, however, have received little scrutiny. Objective.—To evaluate the expression of commonly used immunomarkers and patterns in various acute leukemias to help define the best use and role of multiparameter flow cytometry in the diagnosis and proper classification of acute leukemias. Design.—We have retrospectively analyzed the immunophenotypic data from 508 de novo adult and pediatric acute leukemia patients, as studied using multiparameter flow cytometry in addition to routine morphologic and enzyme cytochemical analysis. Cytogenetic and/or molecular data were correlated in all 41 cases of acute promyelocytic leukemia (APL) and in 203 other cases of acute leukemia where those data were available. We have also determined the positive and negative predictive values of a combined CD34 and HLA-DR expression pattern for the differentiation of APL from other myeloid leukemias. Results.—In acute myeloid leukemia (AML) other than APL, expression of CD34 was seen in 62% and expression of HLA-DR in 86% of all cases. Twenty-six (10%) of 259 cases of non-APL AML were negative for both CD34 and HLA-DR as opposed to 33 (80%) of 41 cases of APL. None of the cases of APL were positive for both CD34 and HLA-DR in contrast to 149 (58%) of 259 cases of non-APL AML. Fifty-three cases were found to be examples of minimally differentiated AML (AML M0) based on the lack of expression of cytoplasmic CD3 and cytoplasmic CD79a and expression of one or more myelomonocytic-associated antigens and/or myeloperoxidase. Expression of CD20 was seen in 40 (24%) of 168 cases of precursor B-cell acute lymphoblastic leukemia (pB-ALL) and 52 (29%) lacked CD34 expression. Five of 180 cases of pB-ALL and 2 cases of precursor T-cell ALL (pT-ALL) were negative for terminal deoxynucleotidyl transferase (TdT). Aside from cytoplasmic CD3, CD5 and CD7 were the most sensitive antigens present in all 21 cases of pT-ALL. CD33 was more sensitive but less specific than CD13 for myeloid lineage. Conclusion.—Aside from identification of blasts, flow cytometry was found to be especially useful in the correct identification of AML M0, differentiation of APL from AML M1/M2, and correct identification of TdT-negative ALL and unusual variants, such as transitional B-cell ALL and undifferentiated and biphenotypic acute leukemias.Keywords:
Immunophenotyping
HLA-DR
Immunophenotyping
Cytometry
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Objective To explore the immunophenotype features of CD34 positive cells in patients with myelodysplastic syndrome(MDS).Methods The immunophenotypes of the CD34 positive cells in bone marrow nuclear cells of 20 patients with MDS were performed with a panel of monoclonal antibodies and four-color flow cytometry by secondary gating strategy.Results With the progression of MDS from RA/RAS,RAEB to RAEBT,the proportion of CD34~+ cells was gradually increased from 7.6%,40.1% to 71.3%(P0.05).Using CD45 vs CD34 gating strategy,the expressions of HLA-DR,CD13,CD33,CD117 on blasts were higher by secondary gating method.However,there were no differences in the expression of above-mentioned antigens on CD34~+ cells in different MDS subtypes(P0.05).Conclusion Secondary gating strategy could accurately reflect the immunophenotype features of blasts in MDS.
Immunophenotyping
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Immunophenotyping
Hematology
Acute lymphocytic leukemia
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Fifty-nine adult patients with acute leukemia were classified using a combination of the French-American-British (FAB) criteria and characterization by immunophenotyping using flow cytometric study. The authors identified 51 patients with acute myeloblastic leukemia and eight with acute lymphoblastic leukemia. This procedure permitted lineage assignment in leukemias that otherwise might have been unclassifiable. In addition, the authors demonstrated that the leukemic blasts of 29% of patients with myeloblastic disease exhibited one or more T-cell antigens on their surface. The use of immunophenotyping has greatly enhanced the authors' ability to correctly identify the lineage of acute leukemias. The data, however, must be interpreted with caution with respect to diagnosing acute mixed lineage leukemias and must be integrated with the morphologic and cytochemical evaluation of traditional classification schemes. The possible significance of T-cell markers in myeloblastic leukemia is discussed.
Immunophenotyping
Acute myeloblastic leukemia
Lineage (genetic)
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Immunophenotyping
Lineage (genetic)
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This study was aimed at exploring the immunophenotypic features of blasts in patients with myelodysplastic syndromes (MDS). Four-color flow cytometry using conventional and secondary gating strategies was used to immunophenotype blasts and the CD34 positive cells in bone marrow nucleated cells of 29 patients with MDS. The results showed: (1) with progression of MDS from RA/RAS, RAEB to RAEB-T, the proportion of CD34(+) cells were gradually increased from 8.0%, 46.4% to 76.8% (P < 0.05); (2) using CD45 vs SSC gating strategy, with the transformation of RA/RAS, RAEB to RAEB-T, the expression of HLA-DR, CD13, CD33, CD117 were also gradually increased (P < 0.05), and the expression of CD15 was gradually decreased (P < 0.05); (3) using CD45 vs CD34 gating strategy, the expression of HLA-DR, CD13, CD33, CD117 on blasts were higher by secondary gating method than those by conventional gating (P < 0.05). However, there were no significant difference (P > 0.05) in the expression of above-mentioned antigens on CD34(+) cells among different MDS subtypes. It is concluded that conventional gating method can reflect MDS progression from RA/RAS, RAEB to RAEB-T, and secondary gating strategy may accurately reflect the biological features of blasts in MDS. Abnormal expression of CD34 is related to the immaturity level and heterogeneity of blast cells, which is beneficial to the diagnosis of clinically suspected MDS incapable of diagnosing with morphology and cytogenetics.
Immunophenotyping
Cytometry
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Data available on request due to privacy/ethical restrictions.
NPM1
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To report on a case of therapy-related acute monocytic leukemia(t-AML) with t(11;17) (q23;q21)/MLL-AF17q after successful treatment for acute promyelocytic leukemia(APL) with t(15;17) (q22;q21)/PML-RARα.A MICM method (bone marrow morphology(M), immunophenotype(I), cytogenetics(C), and molecular biology(M)) was used for the diagnosis and classification of the disease at the time of onset and transformation.The patient was initially identified with typical morphology and immunophenotype of APL. She has carried t(15;17)(q22;q21) and PML-RARα fusion gene but was without t(11;17)(q23;q21) or MLL gene abnormalities. After 13 months of successful treatment, she has transformed to AML with typical morphology and immunophenotype. t(11;17)(q23;q21) and MLL-AF17q fusion gene were detected in her bone marrow sample, while no PLZF-RARα fusion gene was detected by real-time quantitative reverse-transcription PCR(RQ-PCR) and fluorescence in situ hybridization(FISH).t-AML is a serious complication after successful treatment of APL. t(11;17)(q23;q21) is not specific for the diagnosis of variant APL and can also be detected in t-AML. RQ-PCR and FISH are essential for the diagnosis of such patients.
Immunophenotyping
Promyelocytic leukemia protein
Acute monocytic leukemia
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Summary Introduction While the presence of disseminated intravascular coagulation ( DIC ) has been implicated in worse clinical outcome in acute leukemia, the relationship between different subtypes of acute leukemia and the clinicopathologic features of DIC has not been systematically well studied. Methods In this study, we retrospectively reviewed 149 cases of newly diagnosed acute leukemia and assessed the utility of evaluating red blood cell morphologic features, and coagulation parameters in determining the presence of DIC as well as differentiating subtypes of acute leukemia. Results Review of our cohort demonstrates a novel finding, that elevated D‐dimer concentrations ≥19 000 ng/ mL fibrinogen equivalent units ( FEU ) are a sensitive diagnostic indicator of acute promyelocytic leukemia ( APL ) with moderate specificity, sensitivity 96%, specificity 92% in acute leukemia subtyping. Similar to other studies, APL showed an increased incidence of DIC ( P < 0.01) compared to other subtypes of acute leukemia. Surprisingly, the presence of schistocytes on the peripheral blood smear was not a statistically significant indicator of DIC , sensitivity of 36% and specificity of 89%. Finally, the presence of DIC was not a significant indicator of poorer prognosis amongst all patients with AML . Conclusion Overall we identify elevated D‐dimer concentrations ≥19 000 ng/ mL FEU are a sensitive indicator of acute promyelocytic leukemia ( APL ), with a sensitivity of 96% and specificity of 92% in the subtyping of acute leukemias, and that the presence of schistocytes in peripheral blood smears is not a diagnostically sensitive screening test for DIC with a sensitivity of 36%.
D-dimer
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Objective:To investigate the expression of CD 56 on acute leukemia and its clinical manifestation.Methods:Immunophenotype of 92 cases with acute leukemia was measured by flow cytometry,the morphology,immunophenotype and clinic of CD 56 positive patients was analyzed.Results:15/92(16.3%)cases with acute leukemia expressed CD 56 ,one patient has been tesified to be myeloid/NK cell precursor acute leukemia,one patient has been testified to be blastic NK cell leukemia,one patient has been testified to be NK like T cell lymphoma/leukemia,12 patients are like to be acute myeloid leukemia with NK antigen expression or myeloid/NK cell acute leukemia.Conclusion:CD 56 positive acute leukemia is heterogeneous,its morphology,immunophenotype and clinical manifestation is different,they were associated with M 2?M 5?L 2;they had more infiltration out marrow and prognosis was poor.
Immunophenotyping
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