2 SPATIALLY DISTANT EPITOPES OF HUMAN LACTOFERRIN

Lactoferrin (LF) is an iron binding protein, which may represent a tar get for antineutrophil cytoplasmic antibodies (ANCA) in patients affec ted by rheumatoid arthritis, ulcerative colitis, and primary sclerosin g cholangitis, Here we describe the production and characterization of two new monoclonal antibodies (MAbs) against human LF, These MAbs (AG M 10.14, an IgG(1), and AGM 2.29, an IgG(2b)) recognize spatially dist ant epitopes of LF as assessed by cross-blocking experiments, We also demonstrated by indirect immunofluorescence that both MAbs react with ethanol-fixed neutrophil granulocytes showing a perinuclear staining p attern, AGM 2.29 and AGM 10.14 have been utilized as capture and label ed tracer antibody, respectively, in a double determinant immunoassay (DDIA) to measure soluble LF, The results obtained show that this DDIA allows us to quantify even low concentrations of LF, the maximal rang e of the assay sensitivity being between 12 and 780 ng/ml, Therefore, AGM 10.14 and AGM 2.29 may represent useful reagents for studying the role of autoantibodies to LF as well as for measuring soluble LF, whic h is a reliable secretory marker of neutrophil activation.