The flow cytometric detection of aberrant antigen expression is one method proposed for the quantification of minimal residual disease (MRD) in acute leukemias. The present study was designed to investigate the stability of the aberrant antigen expression at relapse or at treatment failure of initial chemotherapy. For this purpose, multiparameter immunophenotyping with a panel of 15 monoclonal antibodies was used at diagnosis as well as at relapse (43 patients with overall 65 aberrations) and at treatment failure (35 patients with overall 66 aberrations). There was a significant decrease in the percentage of the initially described aberrant antigen expression on leukemia blasts at relapse (P = 0.001; n = 65) as well as at treatment failure (P = 0.0001; n = 66) considering all aberrations in the whole leukemia population. Concerning only patients with acute myelogenous leukemia (AML), significant decreases in the aberrant expression could be detected at relapse (P = 0.031; n = 42) and at treatment failure (P = 0.0001; n = 52). The changes in patients with acute lymphoblastic leukemia (ALL) were significant only at relapse (P = 0.006; n = 23). Initially, the most informative aberration was not detectable in four patients at relapse and in seven patients at treatment failure. A decrease of under 50% of the initial value was observed in another 8 patients at relapse and in 10 patients at treatment failure. In further studies assessing the detection of aberrant antigen expression for MRD, quantification of the relapses should be explicitly analyzed regarding the persistence of the initially described aberrant antigen expression. Cytometry (Comm. Clin. Cytometry) 42:247–253, 2000. © 2000 Wiley‐Liss, Inc.

PNH is a disorder of the pluripotent stem cells resulting in a deficient expression of membrane-bound GPI-anchored proteins in different cell types. Several flow cytometric approaches are designed to detect this antigen deficiency. But they all require drawing and testing of normal samples as control. Therefore, in the present study two flow cytometric assays for the detection of CD55 and CD59 deficiency in erythrocytes (REDQUANT CD55/CD59) and granulocytes (CELLQUANT CD55/CD59) are proposed. Precalibrated beads are used to define the cut off between normal and deficient cell populations. The specificity of the tests has been evaluated in healthy blood donors (n=52) resulting in a clear and reproducible cut off (3%) for the normal percentage of GPI-deficient cells. This cut off has been confirmed in leukaemia and lymphoma patients not suspected for developing PNH. The sensitivity has been tested in patients suffering from known PNH (n=23). Both tests performed in combination allowed a reliable detection of PNH in all patients showing antigen deficiencies in both cell types in most patients (20/23). In contrast, the PNH clones in the investigated patients with MDS (4/19) or AA (4/22) were present in granulocytes or erythrocytes, only. This underlines the necessity of analysing erythrocytes as well as granulocytes. Preliminary data regarding a possible correlation between disease activity and percentage of antigen-deficient cells lead to the assumption that haemolytic crises can only be determined on granulocytes whereas deficient erythrocytes disappeared due to complement-mediated lysis of the PNH clone. In conclusion, the combination of the test kits enables the differential diagnosis of PNH clones in a standardized, simple and rapid approach which may have therapeutic consequences.

By simultaneous DNA staining and immunophenotyping, the aneuploid blasts of acute lymphoblastic leukemia (ALL) can be quantitated by flow cytometry. The present study evaluates the application of this method for the detection of minimal residual disease (MRD). For this purpose leukemia cells with known aneuploidy were serially diluted with diploid bone marrow cells obtained after chemotherapy. These mixed samples were immunophenotyped using antibodies CD34, CD10, or CD19 and processed for DNA staining with propidium iodide. Fifty thousand cells were measured for each experiment and the results were compared with the analysis of control sample without added aneuploid cells. Aneuploid cells diluted to 1% were not detectable in 6/15 cases when DNA content analysis was used alone, and in dilutions of < 1%, these cells were not detectable. When DNA content and immunophenotype were simultaneously measured, aneuploidy was detectable in 15/15 cases at 1.00%, 6/6 at 0.50%, 16/16 at 0.10%, and 16/17 at 0.05%. Therefore this method may be useful for the detection of minimal residual disease in cases of aneuploid lymphoblastic leukemia. Cytometry 30:47–53, 1997. © 1997 Wiley‐Liss, Inc.

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