High sensitivity 14-parameter flow cytometry for evaluation of minimal residual disease in childhood B-cell acute lymphoblastic leukemia - a pilot study

Purpose. Minimal residual disease (MRD) is a population of leukemic cells in the bone marrow or in the peripheral blood that is resistant to therapy and leads to relapse. In childhood acute lymphoblastic leukemia (cALL), early and late therapeutic response is assessed by monitoring of MRD. Evaluation of MRD is crucial both for predicting clinical outcomes and for choosing the optimal treatment strategy. The purpose was to evaluate the applicability of new markers for MRD and to define the sensitivity of 14-color flow cytometry (FC) panel. Material/Methods. Materials include preparation of spiked-in samples with defined concentration of leukemic cells by the use of non-leukemic bone marrow aspirates and bone marrow of a child with B-cell precursor ALL (BCP-ALL), rich in blasts. Leukemic cells were added to non-leukemic samples to achieve the following expected blast levels: 1%, 0.1%, 0.01%, 0.001% and 0.0001% and were stained with the newly developed panel. Analysis of FC data was performed by classical manual approach and by automated methods using FlowJo software. Results. Application of the 14 monoclonal antibody panel and the analysis algorithm allow detection of 0.001% blasts, which has a tenfold higher sensitivity than conventional flow cytometry. Conclusion. The newly developed FC panel and analysis algorithm achieve sensitivity similar to that of the genetic methods. This would allow significantly earlier detection of MRD and appropriate therapeutic changes.