title: A simple flow cytometric assay for routine paroxysmal nocturnal hemoglobinuria testing based on immature reticulocytes and granulocytes.
abstract: BACKGROUND
  The aim of this study was to test an easy-to-perform flow cytometric (FCM) assay for the routine investigation for diagnosis of paroxysmal nocturnal hemoglobinuria (PNH), through the simultaneous detection of PNH clones on immature reticulocytes (i-RET) and granulocytes.
  
  
  METHODS
  During the last 5 years, eight patients were diagnosed with PNH in our laboratory, among 90 patients prospectively studied for PNH. The determination of glycosylphosphatidylinositol (GPI) deficient cells on the erythroid lineage was made with a two-color FCM assay of CD71 and CD59, evaluating the PNH clone on i-RET. Three color combinations based on CD66b/CD16/CD45 and CD59/CD24/CD45 were used for the determination of GPI-deficient granulocytes.
  
  
  RESULTS
  In all the patients with PNH, the PNH clones determined with CD71(+)CD59(-) red blood cells (RBC) were nearly identical to the respective clones determined with CD16(dim/-)/CD66b(-) and CD59(-)/CD24(-) granulocytes, in contrast to the clones determined with CD59-deficient erythrocytes only, which were significantly lower.
  
  
  CONCLUSIONS
  Our results indicate that the simultaneous assessment of the PNH clone on CD71(+)/CD59(-)i-RET and CD16(dim/-)/CD66b(-) granulocytes, could offer a reliable method of two series PNH screening, at low cost and with ease of application.
authors.affiliation: Flow Cytometry Laboratory, Department of Immunology, Athens Regional General Hospital G. Gennimatas, Athens, Greece. nikolaostsagarakis@gmail.com
authors.forename: Nikolaos J
authors.initials: NJ
authors.lastname: Tsagarakis
authors.forename: George
authors.initials: G
authors.lastname: Paterakis
mesh_terms: D000328:Adult; D015703:Antigens, CD / Q000097:blood / Q000378:metabolism; D015415:Biomarkers / Q000097:blood / Q000378:metabolism; D005260:Female; D005434:Flow Cytometry; D006457:Hemoglobinuria, Paroxysmal / Q000097:blood / Q000175:diagnosis* / Q000473:pathology; D006801:Humans; D008297:Male; D009504:Neutrophils / Q000378:metabolism*; D011446:Prospective Studies; D012156:Reticulocytes / Q000378:metabolism*
chemical_list: D015703:Antigens, CD; D015415:Biomarkers