title: Multiparameter FLAER-based flow cytometry for screening of paroxysmal nocturnal hemoglobinuria enhances detection rates in patients with aplastic anemia.
abstract: Flow cytometry is the gold standard methodology for screening of paroxysmal nocturnal hemoglobinuria. In the last few years, proaerolysin conjugated with fluorescein (FLAER) has become an important component of antibody panel used for the detection of paroxysmal nocturnal hemoglobinuria (PNH) clone. This study aimed to compare PNH clone detection by flow cytometry in the pre-FLAER era versus the FLAER era. This was a retrospective analysis of 4 years and included 1004 individuals screened for PNH clone, either presenting as hemolytic anemia or as aplastic anemia. In the pre-FLAER time period, the RBCs and neutrophils were screened with antibodies against CD55 and CD59. With the introduction of FLAER, neutrophils were screened with FLAER/CD24/CD15 and monocytes with FLAER/CD14/CD33 combination. A comparative analysis was done for detection of PNH clone in aplastic anemia patients versus non-aplastic anemia patients, as well as between pre-FLAER and FLAER era. Out of a total of 1004 individuals, 59 (5.8%) were detected to have PNH clone positivity. The frequency of PNH clone detected in aplastic anemia and non-aplastic anemia groups was 12.02 and 3.36%, respectively. The detection rate of PNH clone increased from 4.5% (32/711) in the pre-FLAER era to 9.2% (27/293) with the introduction of FLAER. However, this increase could be attributed to increased detection of PNH clone in the aplastic anemia group, which showed a significant increase from 8.3 to 18.2% after use of FLAER. In the non-aplastic group, PNH clone was detected with similar frequencies before and after use of FLAER (3.2 versus 3.8%, respectively). Mean PNH clone size was lower in the aplastic anemia group when compared with the non-aplastic group. RBCs always showed a lower clone size than neutrophils. PNH clone on neutrophils and monocytes was however similar. Inclusion of FLAER increases the sensitivity of the test which is especially useful in picking up small PNH clones in patients of aplastic anemia.
authors.affiliation: Department of Hematology, Postgraduate Institute of Medical Education & Research, Sector-12, Chandigarh, 160012, India.
authors.forename: Man Updesh Singh
authors.initials: MU
authors.lastname: Sachdeva
authors.forename: Neelam
authors.initials: N
authors.lastname: Varma
authors.forename: Dinesh
authors.initials: D
authors.lastname: Chandra
authors.forename: Parveen
authors.initials: P
authors.lastname: Bose
authors.forename: Pankaj
authors.initials: P
authors.lastname: Malhotra
authors.forename: Subhash
authors.initials: S
authors.lastname: Varma
mesh_terms: D000741:Anemia, Aplastic / Q000175:diagnosis*; D004912:Erythrocytes / Q000166:cytology; D005434:Flow Cytometry / Q000379:methods*; D006098:Granulocytes / Q000166:cytology; D006457:Hemoglobinuria, Paroxysmal / Q000175:diagnosis*; D006801:Humans; D012189:Retrospective Studies