A sensitive time-resolved immunofluorometric assay of ferritin in serum with monoclonal antibodies

We describe a new non-isotopic immunoassay for the quantitation of ferritin in serum. Ferritin is first immunoextracted from serum with a monoclonal antibody immobilized in white microtiter wells. A second biotinylated antibody is then added. The bound biotinylated antibody is quantified by a bridge reaction with streptavidin labeled with the europium chelate 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid (BCPDA) in the presence of excess Eu3+. The fluorescence on the bottom of the dry microtiter well consisting of monoclonal antibody-ferritin-monoclonal antibody-biotin-streptavidin-BCPDA-Eu3+ is then measured with a time-resolved fluorometer. The assay is sensitive (0.5 microgram/l), precise (CV's 3-9%) and accurate (average recovery 102%). Results compared well (r = 0.99) with those of a widely used RIA procedure.