(A) Strategy applied for flow cytometric AML MRD multicenter harmonization by the ALFA Intergroup. [1] Rationale of AML MRD flow panel design was based on simplicity, reproducibility, and cost. Tube 1 was a core combination for LAIP detected at diagnosis and/or by different-from-normal analysis, tube 2 was targeted to aberrancies of CD34⁺CD38⁻ cells (immunophenotypic LSCs), and tube 3 was an optional development tube for monocytic aberrancies. [2] Flow cytometer fluorescent settings were harmonized (“mirrored”) between Canto vs Navios cytometer platforms. Voltages were set to reach target mean fluorescence intensity (MFI) values by acquisition of rainbow calibration beads without compensation for fluorescent channels FL1 to FL8 on the Canto cytometers; these rainbow bead settings were transposed to Navios cytometers by applying MFI target = Canto target/256. Mirrored (superimposable) target peaks for both cytometers are shown for FL1 and FL8 fluorescent channels with an example of resulting comparable antibody profiles between cytometer platforms. (B) [3] Quality controls for reproducibility of staining profiles from harmonized cytometer settings/sample processing between cytometers/laboratories. Examples shown are for CD117 and CD38 expression intensity on CD34⁺ gated mononuclear cells of tube 1 from 10 shared BM samples stained and then acquired on Canto or Navios flow cytometers. Intensity profiles are similar between cytometer platforms for each sample. [4] External quality assessment for all harmonized steps from preanalytical to final gating analyses by distribution of a normal BM to 22 participating laboratories (cytometer platforms: 12 Cantos, 10 Navios). Example shows that strong reproducibility can be achieved in the detection of rare events (shown for CD34⁺CD38⁻) among 22 participating laboratories. Figure by Christophe Roumier and Adriana Plesa, ALFA.