The use of monoclonal antibodies and flow cytometry in the diagnosis of paroxysmal nocturnal hemoglobinuria

Abstract

We have characterized the erythrocytes, granulocytes, and platelets of 54 patients with paroxysmal nocturnal hemoglobinuria (PNH) with antibodies to glycosylphosphatidylinositol-anchored proteins (anti- CD55, anti-CD59, and anti-CD16) and flow cytometry to establish the usefulness of this technique in the diagnosis of this disorder. All patients demonstrated either completely (PNH III) or partially (PNH II) deficient red cells and granulocytes. Anti-CD59 best demonstrated PNH II red cells, which were present in 50% of the patients. The proportion of abnormal granulocytes was usually greater than the proportion of abnormal red cells; 37% of the patients had >80% abnormal granulocytes. Anti-CD55 did not delineate the erythrocyte populations as well as did anti-CD59. Either anti-CD55 or anti-CD59 could be used equally well to analyze granulocytes; anti-CD16 did not demonstrate cells of partial deficiency. Platelets could not be used for detailed analysis as the normal and abnormal populations were not well distinguished. Flow cytometry of erythrocytes using anti-CD59 or of granulocytes using either anti-CD55 or anti-CD59 provides the most accurate technique for the diagnosis of paroxysmal nocturnal hemoglobinuria; it is clearly more specific, more quantitative, and more sensitive than the tests for PNH that depend upon hemolysis by complement (the acidified serum lysis [Ham] test, the sucrose lysis test, and the complement lysis sensitivity [CLS] test).

Changing prognosis in paroxysmal nocturnal haemoglobinuria disease subcategories: an analysis of the International PNH Registry

“…Paroxysmal nocturnal haemoglobinuria (PNH) is a rare, acquired disease of the haemopoietic stem cell, characterised by chronic intravascular haemolysis and by marrow failure. [1][2][3][4][5][6] The cellular abnormality in this life-threatening disease originates from a mutation in the phosphatidylinositol glycan class A (PIGA) gene, resulting in a deficiency of glycosylphosphatidyl-inositol (GPI)-anchored complement regulatory proteins, including CD55 and CD59, from the surface of blood cells. 7 Patients with PNH haemolysis experience a marked increased risk of thromboembolism (TE), which is the leading cause of death.…”

Flow cytometric study of neutrophilic granulopoiesis in normal bone marrow using an expanded panel of antibodies: Correlation with morphologic assessments

“…Early studies of granulocytic surface antigens focused on the diagnosis of certain inherited disorders, such as leukocyte adhesion molecule deficiency, in which a specific antigen is known to be missing (1). Studies of peripheral blood (PB) granulocytes have expanded to include other antigens, such as CD55 and CD59, for the diagnosis of paroxysmal nocturnal hemoglobinuria (2). Since these studies are performed on PB, they may not pose a great challenge because the vast majority of PB granulocytes are of the same maturation stage (i.e., segmented neutrophils).…”

Multilineage glycosylphosphatidylinositol anchor‐deficient haematopoiesis in untreated aplastic anaemia

“…The phenotypic hallmark of PNH blood cells is a deficiency of cell surface glycosylphatidylinositol (GPI)anchored proteins (Hall & Rosse, 1996;Rosse, 1997), owing to mutation of the PIGA gene that encodes the enzyme required for the first step in the biosynthesis of the GPI anchor itself. Using monoclonal antibodies directed against individual GPI-anchored proteins, PNH cells have been detected in .…”