An aptamer-based four-color fluorometic method for simultaneous determination and imaging of alpha-fetoprotein, vascular endothelial growth factor-165, carcinoembryonic antigen and human epidermal growth factor receptor 2 in living cells.

Abstract

The extraordinary fluorescence quenching capability of graphene oxide (GO) was coupled to the specific recognition capability of aptamers to design a four-color fluorescent nanoprobe for multiplexed detection and imaging of tumor-associated proteins in living cells. Specifically, alpha-fetoprotein (AFP), vascular endothelial growth factor-165 (VEGF165), carcinoembryonic antigen (CEA), and human epidermal growth factor receptor 2 (HER2) were detected. Due to strong π interaction, the fluorescence of labeled aptamers is quenched by GO. Four fluorophore-labeled aptamers that bind the tumor-associated proteins were adsorbed on GO to form the four-color nanoprobe with quenched fluorescence. The nanoprobes were internalized into cells via endocytosis, where the aptamer/GO nanoprobes bind the intracellular tumor-associated proteins. The aptamer-protein complexes thus formed detach from GO, and fluorescence recovers. Each analyte has its typical color (AFP: blue; VEGF165: green; CEA: yellow; HER2: red). As a result, simultaneous detection and imaging of multiple tumor-associated proteins in living cells were achieved. This nanoprobe has a fast response and is highly specific and biocompatible. The linear ranges for AFP, VEGF165, CEA, and HER2 are 0.8nM-160nM, 0.5nM-100nM, 1.0nM-200nM, and 1.2nM-240nM, respectively. Detection limits were 0.45nM for AFP, 0.30nM for VEGF165, 0.62nM for CEA, and 0.96nM for HER2. The probe allows for a fast distinction between tumor cells and normal cells via imaging. Graphical abstract Schematic presentation of the development of a four-color fluorometic method based on aptamer and graphene oxide for simultaneous detection and imaging of alpha-fetoprotein, vascular endothelial growth factor-165, carcinoembryonic antigen and human epidermal growth factor receptor 2 in living cells.