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

Abstract

A liquid-phase immunoassay was developed for the simultaneous determination of five cancer biomarker proteins: alpha-fetoprotein (AFP), human chorionic gonadotropin (hCG), carcinoembryonic antigen (CEA), ovarian tumor antigen (CA125/MUC16), and gastrointestinal tumor antigen (CA19-9). The method was based on the incubation of a serum (or tissue cytosol) with five antibodies, each labeled with a different lanthanide (Pr(3+), Eu(3+), Gd(3+), Ho(3+), and Tb(3+), 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 the 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.

Magnetic immunoassay coupled with inductively coupled plasma mass spectrometry for simultaneous quantification of alpha-fetoprotein and carcinoembryonic antigen in human serum

“…Table 3 shows the comparison of the proposed method with other methods for the analysis of AFP and CEA. It can be seen that the established method exhibited lower or comparable LODs than that obtained by other ICP-MS based immunoassay method [20,37,35,38] and higher than that obtained by the ICP-MS-based method involving single particle detection mode [23]. Besides, the established magnetic immunoassay method allowed simultaneous determination of AFP and CEA and showed higher sensitivity over chemiluminescent immunoassay.…”

Aptamer-based microchip electrophoresis assays for amplification detection of carcinoembryonic antigen

“…This LOD value was far below the threshold of CEA (2.5 ng/ml) in normal human serum [32]. It was also below the LOD values obtained from inductively coupled plasma mass spectrometry (0.14 ng/ml) [33], capillary-based three-dimensional immunosensor (0.2 ng/ml) [34], size exclusion chromatography with inductively coupled plasma mass spectrometric detection (2.6 ng/ml) [35], rolling circle amplification for protein detection (6.8×10 −12 M) [11], and isothermal circular strand-displacement polymerization for protein detection (12 ng/ml) [13]. Assay reproducibility was estimated by analyzing a standard solution of CEA at 5.0 ng/ml (n=7).…”

A Lanthanide-Based Chemosensor for Bioavailable Fe3+ Using a Fluorescent Siderophore: An Assay Displacement Approach

“…β-Diketonate ligands coordinate via the two oxygen atoms leading to neutral species in a 3:1 β-diketonate:lanthanide ratio, the complexes have been shown to be stable in aqueous solutions [35]. Therefore these molecules have been used in many applications from sensing [36], antibody labeling [37], new materials such as liquid crystals [38], near IR-LEDs, [39] sol-gel glasses [40], and polymers [40]. …”