Solid-phase competitive assay utilizing a fusion protein

摘要

An in vitro solid-phase, competitive assay for detecting the presence of a peptide analyte in a biological sample comprises contacting a fusion protein, of first and second peptides, with a solid supported first antibody or fragment thereof, which binds the first peptide, adding a biological sample containing the peptide analyte and a second antibody or fragment thereof which binds the peptide analyte and the second peptide. The second antibody is allowed to bind any free analyte present in the sample, and to form solid supported complexes of fusion protein and antibody. The amount of solid supported second antibody detected is then compared to a control. When the amount of peptide analyte in the sample increases the amount of second antibody bound to the solid support decreases. An optional final addition of an antibody-binding molecule helps detect the solid supported complexes of the fusion protein and the second antibody or fragment. The assay may be utilized for detecting neoplastic antigens from tumors or their metastases by using a second peptide that binds specifically to an antibody specific to an antigen, such as an epithelial mammary cell antigen. A kit for detecting the presence of neoplastic tissue comprises the fusion protein, a first antibody or fragment thereof specifically binding the first peptide, and a second antibody or fragment thereof specifically binding the second peptide, and optionally a device for detecting the second antibody or fragment thereof, such as a labeled antibody-binding molecule.