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PNH with FLAER High Sensitivity Test codes 16433. Question 1. What are the clinical and genetic features of paroxysmal nocturnal hemoglobinuria PNH? Paroxysmal nocturnal hemoglobinuria PNH is a rare disorder characterized by chronic hemolysis, increased risk. Neut – FLAER / CD24 / CD66b. Mono – FLAER / CD14. Then if PNH clone found, test red cells to identify nature of deficiency Type 2 vs 3 2 Stages. Use CD235a Glycophorin A to identify red cell pop. not GPI-linked Then use GPI-linked marker to find deficient pop. PNH cells do not produce the anchor, therefore FLAER will not bind to these cells. Before FLAER, detection of PNH clones by flow cytometry relied on fluorescently labeled antibodies to GPI-linked proteins such as CD59 and CD55. Unfortunately, these antibodies do not bind with high affinity to their ligands, so that small PNH clones were not. PNH clones were detected with the FLAER assay in 63 11.8% of 536 samples tested, whereas PNH RBCs were detected in only 33 6.2%, and always with a smaller clone size. The FLAER assay on WBCs is a more sensitive and robust primary screening assay for detecting PNH clones in. A, Fresh PNH sample stained with FLAER, CD33, CD45, and CD14. Of monocytes from R2, 72.7% exhibit a CD14–/FLAER– PNH phenotype lower left; 81% of neutrophils from R3 exhibit a CD14–/FLAER– PNH phenotype lower middle. About 14% of the lymphocytes in this sample were also unstained by FLAER lower right.

Paroxysmal Nocturnal Hemoglobinuria PNH, Flow Cytometry;. FLAER is a fluorescently labeled variant of aerolysin that binds directly to the GPI anchor and can be used to evaluate the expression of the GPI linkage. Individuals with PNH have decreased or absent expression of CD14 on monocytes. What is Paroxysmal Nocturnal Haemoglobinuria. PNH is a rare blood condition where blood cells are prone to be attacked by part of the body’s immune system. The process where the red blood cells are destroyed is called “haemolysis” and it is responsible for many of the symptoms of the disease. Before FLAER, detection of PNH clones by flowcytometry relied on fluo-rescently labeled antibodies to GPI-linked proteins such as CD59 and CD55. These antibodies do not bind with high affinity,so that small PNH clones are not detected. Also, they each screen for the absence of a specificprotein. FLAER is an Alexa Fluor® 488 labeled variant of aerolysin, a unique protein that binds tightly and specifically to mammalian GPI anchors. FLAER will not bind to PNH cells because they do not produce the anchor. Before FLAER, detection of PNH clones by flow cytometry relied on fluorescently labeled antibodies to GPI-linked.

106 Detection of PNH cells by flow cytometry, using multiparameter analysis Flow cytometry is nowadays considered the methodology of choice for detection of PNH clones3, 15. eseguito l’approfondimento diagnostico per PNH 1. L’analisi citometrica evidenzia la presenza di un clone compatibile con PNH pari al 80% sui granulociti, al 76% sui monociti e al 14% sugli eritrociti. In aggiunta a FLAER, gli antigeni “GPI-linked” valutati sono CD66b sui granulociti identificati mediante “gate” CD45/SSC.

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