The authors report on the conjugation of monoclonal antibodies against the biomarker epithelial cell adhesion molecule (EpCAM) to silica nanoparticles doped with the dye Cy5 (Cy5-SiNPs). Conjugation was performed on the Cy5-SiNPs that were previously coated with a layer of protein G which serves as a linker controlling the orientation of the antibody. The conjugation method takes advantage of site specific interactions between the protein G and constant domains (Fc) of the antibody. The method warrants the antibody binding sites (Fab) to be faced outwards such that the conjugates maintain their affinity for binding the analyte (EpCAM). In vitro analysis by confocal fluorescence imaging and flow cytometry using analytical wavelengths comparable with the excitation and emission wavelength of Cy5-SiNPs at 643 and 662 nm, respectively. The result demonstrated the oriented conjugate to specifically bind to target cells (HT-29) with a sensitivity that is 12 times higher than that of conjugates prepared by conventional EDC coupling. In vivo fluorescence imaging of mice bearing the HT-29 tumor highlighted time-dependent accumulation of the oriented conjugates at the tumor site. As indicated by biodistribution studies hepatic excretion of the oriented probes occurs, however tumor fluorescence still remains for up to 14 days post injection. This research demonstrates that the oriented conjugates derived herein can improve target cell detection sensitivity and can be successfully applied in tumor imaging, which should drive further development of new classes of effective fluorescence contrast agents for cancer diagnostics.
Graphical abstract Cy5 dye-doped silica nanoparticles were conjugated to antibodies specific for the epithelial cell adhesion molecule. The nanoparticles were previously coated with protein G to control the orientation of the antibody. This warrants enhanced sensitivity for in vitro analysis and also enables in vivo imaging.
The authors describe a highly sensitive and selective photoelectrochemical (PEC) assay for mercury(II) ions. It is based on a dual signal amplification strategy. The first enhancement results from the surface plasmon resonance (SPR) of Au@Ag nanoparticles (NPs) absorbed on MoS2 nanosheets. Here, the injection of hot electrons of Au@Ag NPs into MoS2 nanosheets produces a strong photocurrent, while background signals are strongly reduced. The second enhancement results from the use of a thymine rich ct-DNA aptamer attached to the Au@Ag-MoS2 nanohybrid. The DNA specifically binds Hg(II) ions to form thymine-Hg(II)-thymine (T-Hg-T) complexes. This leads to the formation of a hairpin-shaped dsDNA structure. The use of a CdSe quantum dot label at the terminal end of the ct-DNA further facilitates electron–hole separation. The photocurrent of the detector is measured as a function of Hg(II) concentration at a bias voltage of 0.1 V and under irradiation of 430 nm light. Due to the two-fold amplification strategy presented here, the linear range extends from 10 pmol·L?1</sup> to 100 nmol·L?1</sup>, with a detection limit of 5 pmol·L?1</sup> (at S/N?=?3).
Graphical Abstract The injection of hot electrons of Au@Ag into MoS2 produces a strong photocurrent, and the formation of thymine-Hg(II)-thymine further facilitates electron–hole separation by CdSe. This dual signal amplification strategy is used to detect Hg(II) ions via a photoelectrochemical assay.
