Multiplexed Determination of Protein Biomarkers Using Metal-Tagged Antibodies and Size Exclusion Chromatography-Inductively Coupled Plasma Mass Spectrometry

A liquid-phase immunoassay was developed for the simultaneous determination of five cancer biomarker proteins: alpha-fetoprotein (AFP), human chorionic gonadotropin (KG), carcinoembryonic antigen (CEA), ovarian tumor antigen (CA125/MUC16), and gastrointestinal tumor antigen (CA199). The method was based on the incubation of a serum (or tissue cytosol) with five antibodies, each labeled with a different lanthanide (Pr3+, Eu3+, Gd3+, Ho3+, and Th3+, respectively) followed by the specific determination of the immunocomplex formed by size exclusion chromatography with inductively coupled plasma mass spectrometric detection (SEC-ICPMS). The sensitivity of the method was comparable with that attainable by enzyme-linked immunosorbent assay (ELISA) or radioimmunoassay with die advantages of multiplexed analysis capacity, virtually no sample preparation, and sample amount consumption, ca. 3 times lower than an ELISA test. The method was validated for the analysis of the proteins in human serum and proved to be able to discriminate ovary and uterus tumor tissue samples from those of healthy subjects.