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Clinical evaluation of an aerolysin‐based screening test for paroxysmal nocturnal haemoglobinuria
Author(s) -
Peghini Pietro Edmondo,
Fehr Jörg
Publication year - 2005
Publication title -
cytometry part b: clinical cytometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.646
H-Index - 61
eISSN - 1552-4957
pISSN - 1552-4949
DOI - 10.1002/cyto.b.20059
Subject(s) - antibody , haemolysis , microbiology and biotechnology , staining , paroxysmal nocturnal hemoglobinuria , chemistry , immunology , medicine , biology , pathology
Background Recently, a toxin produced by Aeromonas hydrophila was demonstrated to bind directly to the glycosyl‐phosphatidyl‐inositol (GPI) anchor. After coupling it to a fluorescent dye and applying it in fluorescence‐activated cell scanning (FACS), this property was exploited to detect GPI‐negative cells in the diagnosis of paroxysmal nocturnal haemoglobinuria (PNH). Methods We used this reagent according to a very simple staining protocol followed by single‐colour FACS and compared the results in patients with PNH and normal controls with those obtained with antibody‐mediated detection of cells lacking GPI‐anchored proteins. Results We observed very good concordance between the two methods, with correlation coefficients (R 2 ) of quantified GPI‐deficient cell populations ranging from 0.952 to 0.969. The lower limit of detection was determined at 0.50% GPI‐negative cells, which was in the range obtained with double‐colour staining with antibodies (0.20–1.00%, depending on the antibody). A significant correlation was observed between the fraction of GPI‐negative granulocytes and laboratory parameters of haemolysis, with the erythrocyte creatine having the best correlation (R 2 = 0.671, P < 0.0001). Conclusions Using this protocol, we were able to reliably diagnose PNH with a high sensitivity. The test allows the identification of GPI‐negative granulocyte populations as small as 0.5%. © 2005 Wiley‐Liss, Inc.