Standardized high-sensitivity flow cytometry testing for paroxysmal nocturnal hemoglobinuria in children with acquired bone marrow failure disorders: A single center US study

Cytometry B Clin Cytom. 2018 Jul;94(4):699-704. doi: 10.1002/cyto.b.21536. Epub 2017 Jun 27.


Background: Paroxysmal nocturnal hemoglobinuria (PNH) is a rare, acquired hematopoietic stem cell disorder that has not been well-documented in children, particularly those with acquired bone marrow failure disorders (ABMFD)-acquired aplastic anemia (AAA) and myelodysplastic syndrome (MDS). Therefore, we sought to determine the prevalence of PNH populations in children with ABMFD.

Methods: PNH testing was performed in children with an ABMFD diagnosis using high sensitivity (≥0.01%) fluorescent aerolysin (FLAER)-based assay according to 2010 International Clinical Cytometry Society (ICCS) PNH Consensus Guidelines and 2012 Practical PNH Guidelines. FLAER/CD64/CD15/CD24/CD14/CD45 and CD235a/CD59 panels were used for white blood cell and red blood cell testing, respectively.

Results: Thirty-seven patients with ABMFD (34 AAA, 3 MDS) were included (17M/20F, age 2-18 years, median 9 years). PNH populations were identified in 17 of 37 (46%) patients. Of the 17 patients with PNH populations identified, 7 were PNH clones (>1% PNH population), and 10 had minor PNH population or rare cells with PNH phenotype (≤1% PNH population).

Conclusions: This is the first study to use a standardized high-sensitivity FLAER-based flow cytometry assay and the recommended cutoff of 0.01% to identify cells with PNH phenotype in pediatric patients with ABMFD in the United States. The identification of a PNH population in 46% of ABMFD supports the recommendation for high sensitivity PNH testing in children with these disorders. As a less sensitive assay using a cutoff of ≥ 1% PNH population would have missed 10 (27%) patients with minor PNH population or rare cells with PNH phenotype. © 2017 International Clinical Cytometry Society.

Keywords: FLAER; ICCS guidelines; acquire bone marrow failure; aplastic anemia (AA); children; flow cytometry; high-sensitivity; myelodysplastic syndrome (MDS); paroxysmal nocturnal hemoglobinuria (PNH); pediatric.

MeSH terms

  • Adolescent
  • Anemia, Aplastic / complications*
  • Child
  • Child, Preschool
  • Female
  • Flow Cytometry / methods*
  • Flow Cytometry / standards
  • Hemoglobinuria, Paroxysmal / diagnosis*
  • Hemoglobinuria, Paroxysmal / epidemiology*
  • Humans
  • Male
  • Myelodysplastic Syndromes / complications*
  • Prevalence
  • Retrospective Studies
  • Sensitivity and Specificity