Comparison of Nonsequencing Techniques for Identification of NPM1 Mutations and Associated Blast Morphology in Patients With Acute Myeloid Leukemia

center dot Context.-Nucleophosmin 1 (NPM1) mutations affect 20% to 30% of all acute myeloid leukemia (AML) patients; several methods are employed to analyze NPM1 muta-tions, each of them with its advantages and limitations.Objective.-To compare 3 nonsequencing protocols capable of detecting the main NPM1 mutations and to evaluate nuclear morphometric analysis (NMA) as an alternative to cuplike blast detection.Design.-We selected multiparameter flow cytometry (MFC), amplification refractory mutation system-polymer-ase chain reaction (ARMS-PCR), and a quantitative PCR (qPCR) kit to identify NPM1 mutations in AML patients at diagnosis. We also evaluated the presence of cuplike blasts and assessed nuclear morphometry using NMA.Results.-MFC appears as a screening method for NPM1 mutations because of its lower specificity. ARMS-PCR demonstrated specificity similar to that of the qPCR kit, although it was more laborious. qPCR testing, conversely, is relatively fast and easy to standardize. Of these methods, qPCR was the only one capable of identifying the type of NPM1 mutation. With regard to morphology, NMA could be used as an alternative for the evaluation of cuplike blasts in AML smears.Conclusions.-qPCR appears to be the best option to identify NPM1 mutations, with ARMS-PCR representing a cheaper alternative. MFC may be used as a screening method, in which results falling within and above the gray zone should be confirmed by molecular testing.(Arch Pathol Lab Med. 2023;147:701-709; doi: 10.5858/ arpa.2021-0601-OA)