Demonstration of cytoplasmic and nuclear antigens in acute leukaemia using flow cytometry.

Abstract

Aims-To detect cytoplasmic and nuclear antigens using flow cytometry in acute leukaemia and to use this technique for double marker combinations.Methods-Cytoplasmic staining was carried out in samples from 40 cases of acute leukaemia with monoclonal antibodies against the myeloid antigen CD13, the lymphoid antigens CD3, CD22, p chain and the enzymes terminal deoxynucleotidyl transferase (TdT) and myeloperoxidase (MPO We describe a technique that does not disrupt the cells and preserves both antigen expression and light scatter, and permits a precise study of the leukaemic cells by gating the desired population for analysis. MethodsPeripheral blood and bone marrow anticoagulated samples from 40 patients with acute leukaemia at presentation or relapse were studied. The diagnosis was based on morphology and cytochemistry according to the FAB criteria.8 9

Myeloperoxidase Detection by Three-Color Flow Cytometry in Acute Lymphoblastic Leukemia

“…They analyzed 4 cases of B-precursor acute lymphoblastic leukemia (ALL) and all were negative for myeloperoxidase (MPO) by enzyme cytochemistry and by flow cytometry (using the Caltag's Fix and Perm Cell Permeabilizing kit). This is in contrast to previous reports by Farahat et al 2 and Hoyer et al 3 in which flow cytometric detection of MPO was described in 7 of 10 cases and in 1 ALL case respectively. They postulate that the different results may be due to the different lysing agents used to permeabilize the cell membrane, and they consider that a larger study of B-precursor ALL cases using Caltag's lysing reagent might be useful in determining the effects of the different permeabilization methods in detecting MPO reactivity in lymphoblasts.…”

Myeloperoxidase Detection by Three-Color Flow Cytometry and by Enzyme Cytochemistry in the Classification of Acute Leukemia

“…In the study by Farahat et al 9 of 33 cases of acute leukemia that were studied for MPO expression by flow cytometry and by the alkaline phosphatase-anti-alkaline phosphatase method, all 3 cases that were negative by flow cytometry were also negative by the alkaline phosphatase-anti-alkaline phosphatase method. Considered together, these results suggest that a negative MPO result by flow cytometry can be considered conclusive evidence of a lack of MPO expression.…”

“…However, despite several series reporting their use, [3][4][5][6][7][8][9][10][11] no large series has compared detection of MPO by enzyme cytochemistry-the FAB "gold standard"-with flow cytometry. Among the larger series, that by Farahat et al 9 We compared conventional enzyme cytochemistry and 3-color flow cytometry in the detection of MPO in a large group of acute leukemias. The aims of the study were to determine whether flow cytometric detection of MPO correlates with the results obtained by cytochemistry and to describe the pattern of MPO expression as detected by flow cytometry in the different FAB subtypes of acute leukemia.…”

Chromosome 17 abnormalities and mutation of the TP53 gene: correlation between cytogenetics, flow cytometry and molecular analysis in three cases of chronic myeloid leukemia

“…For flow cytometry, the samples were fixed and permeabilized as described by Faharat et al (1994) and then incubated for 30 min with 10 mL of anti-p53 MoAb labeled with FITC, followed by two washes with Tween 20/PBS. The cells were then resuspended in 500 mL of PBS/formaldehyde prior to analysis.…”