Flow cytometry thresholds of myeloperoxidase detection to discriminate between acute lymphoblastic or myeloblastic leukaemia

Abstract

SummaryThe World Health Organization 2008 Classification emphasizes myeloperoxidase (MPO) detection as sufficient for assigning a blast population to the myeloid lineage. Published MPO positivity thresholds are 10% for flow cytometry (FCM) but 3% for cytochemistry. Here we re-evaluated the FCM-MPO threshold by comparing retrospectively 128 acute lymphoblastic leukaemias and 75 acute myeloid leukaemias without maturation, all assessed by benzidine-based cytochemistry. A 13% threshold was found to be relevant using an isotype control as background-reference (sensitivity 95Á1%, specificity 91Á7%). Residual normal lymphocytes proved to be an advantageous alternative reference, a threshold of 28% yielding improved 97Á4% sensitivity and 96Á1% specificity.

B-Acute Lymphoblastic Leukemia/Lymphoblastic Lymphoma

“…96 MPO detection can be performed by cytochemistry, FC, and/or immunohistochemistry (IHC), and 3% and 10% cutoffs have been used historically as thresholds for MPO positivity by cytochemistry and FC, respectively. 97 In a recent study, Guy et al 99 proposed a refined threshold of 13% or 28% MPO positivity (using either isotype controls or normal lymphocytes as negative controls) to establish myeloid differentiation by FC immunophenotyping. However, MPO stains by either IHC or FC can be difficult to quantify and are prone to technical artifacts.…”

Acute Leukemias of Ambiguous Origin

“…In a study comparing MPO expression detected by FCM and cytochemistry in cases of AML and ALL, a 13% threshold was found to be relevant using an isotype control as a background reference (sensitivity, 95.1%; specificity, 91.7%). 5 If residual normal lymphocytes were used as reference, a threshold of 28% had to be applied, yielding an improved 97.4% sensitivity and 96.1% specificity in distinguishing between ALL and AML. The WHO criteria for MPAL do not indicate any lower limit for MPO expression in leukemic blasts, 1 but it seems reasonable to use published thresholds.…”

“…The WHO criteria for MPAL do not indicate any lower limit for MPO expression in leukemic blasts, 1 but it seems reasonable to use published thresholds. 3,5 Since MPAL cases often show more than one population of blasts, MPO could be present in only a minor population, which has to be identified as the myeloid component. Another issue is that AML with minimal differentiation and no MPO expression could be involved in MPAL.…”

“…MPO stands as the most robust marker that will identify myeloid engagement. Recent publications have redefined the threshold for MPO positivity, 5,6 and yet most MPAL cases with a myeloid component will express MPO broadly in most blasts. An alternative, for MPAL with a strong monocytic differentiation, is represented by (very rare) cases in which blasts will be positive for nonspecific esterase and/or express at least two of the following antigens: CD14, CD11c, CD36, CD64, or cytoplasmic lysozyme.…”

Haematological profile of patients with mixed-phenotype acute leukaemia from a tertiary care centre of north India

“…However, no single cut-off percentage has been uniformly accepted. Literature reports the use of a wide variety of cut-off percentages, ranging from 3 to 20 per cent in different studies28. Since defining MPO positivity on flow cytometry is often critical for identification of MPAL, each flow laboratory needs to carefully standardize sample processing.…”