Double immunoenzymatic APAAP staining for the detection of leukemia-associated immunophenotypes

Detection of unusual or aberrant cell immunophenotype with flow cytometry i s the basis for the immunologic recognition of minimal residual disease (MR D) in patients with acute leukemia (AL). In this study, we have shown that the double immunocytochemical alkaline phosphatase antialkaline phosphatase (APAAP) staining technique also makes possible the detection of leukemic c ells with unusual (leukemic) combinations of antigens (ULCA) both at diagno sis and during follow-up of patients with ULCA(+) AL. The applicability of double APAAP was analyzed on bone marrow (BM) samples obtained from 12 pati ents (8 with AML, 3 with ALL, and 1 with undifferentiated acute leukemia [A UL]) randomly chosen from a larger group of 22 ULCA(+) patients treated at our center in a 3-year period (22% observed ULCA(+) AL frequency). The perc entages of ULCA(+) BM cells before chemotherapy were in the range of 5%-60% , which dropped to 0%-7% in 10 patients who achieved remission (range 0%-7% , p < 0.01). However, these cells could also be found 60 days after the ini tiation of therapy, ranging from 0%-2% of all nucleated cells. In 2 of 10 p atients who achieved remission, 2% ULCA(+) BM cells were found on days 35 a nd 60 after initiation of chemotherapy, and this finding was followed by re lapse on days 110 and 270. However, the other 8 patients remained in remiss ion despite positive finding of ULCA(+) BM cells ranging from 0.2%-2% on at least one occasion. In 2 patients with AML FAB-M3 and cytomorphologic remi ssion, the finding of ULCA(+) cells by double APAAP correlated with the mol ecular finding of PML/RAR alpha junction. These results indicate that doubl e APAAP staining can identify leukemic cells in samples with a cytomorpholo gic pattern consistent with remission, but its applicability in detection o f MRD awaits additional studies on a larger number of patients with ULCA(+) AL.