Established Standards of Minimal Residual Disease Detected By Multiparametric Flow Cytometry in Acute Leukemia

Purpose

To establish a method for detecting minimal residual disease (MRD) by eight color flow cytometry which is stable, repeatable and accurate quantitation.

Method

According to the ratio of 10%, 1%, 0.1%, 0.01% and 0.001%,to analyze sensitivity of the method by successively mixing the cell lines (Kasumi, KG-1a) and primary acute leukemic bone marrow cell were mixed with normal bone marrow cells. In order to ensure the specificity of the test results, we increased antibody number to eight color combinations of antibodies to adjust fluorescence compensation value after labeling antibody separately in each channel. To verify the feasibility of standardization, standardized test were in 25 bone marrow samples of acute leukemia.

Result

In standard conditions of detection and sensitivity of 10^-5could be detected by eight color flow cytometry. In our study there were 25 cases of acute leukemia, including 14 patients with acute myeloid leukemia and 11 patients with acute B-lymphoblastic leukemia. 23 of 25 cases were detected specific leukemia associated immunophenotypes (LAIP) at diagnosis. 20 patients could be detected the original LAIP, and LAIP of 3 patients changed after remission induction therapy. To analyze the relationship between the clinical data and MRD level, the result showed that the type of LAIP had significant influence on level of MRD. After remission the level of MRD in patients who expressed cross lineage and non-synchronous antigens at diagnosis was significantly higher than those who did not express (P=0.003, P=0.006).

Conclusion

We established the standardized conditions of minimal residual disease detected by multiparametric flow cytometry to ensure the accuracy and specificity of the test results. It has important significance to confirm that manifestations of LAIP were consistent with the outcome in patients with acute leukemia.