Highly sensitive electrochemical immunoassay integrated with polymeric nanocomposites and enhanced SiO2@Au core-shell nanobioprobes for SirT1 determination.

An ultra-highly sensitive electrochemical immunosensor for SirT1 (a key protein in age-related diseases) evaluation has been designed, employing polymeric nanocomposites as sensing platform and core-shell SiO2@Au to immobilize HRP-Ab2 as nanobioprobes. The approach includes chemical synthesis of PAMAM-Au-MWCNT nanocomposites (PNCs) with abundant PAMAM-Au nanoparticles immobilized on the MWCNT matrix, and biochemically synthesis of nanobioprobes with highly dispersed SiO2@Au tracing tags for successive efficient load functionalized enzyme-antibodies (HRP-Ab2). The PNCs nanocomposites could improve the efficiency of immune response via abundant capture antibodies (Ab1#1) on the electrode surface, and accelerate electron transfer through MWCNT and Au nanoparticles. Besides, the SiO2@Au was employed as tracing tags to label numerous HRP-Ab2 to further enhance signal readout during HRP-thionine-H2O2 system. Under optimal conditions, the signal intensity was linearly related to the concentration of SirT1 in the range of 20 pg mL-1 to 500 ng ml-1, and the limit of detection was 12.5 pg mL-1. It is noteworthy that the proposed immunoassay protocol has been successfully applied to evaluate SirT1 expression in cells by different treatment with high sensitivity and accuracy.

A TiO2-Au-polymer hybrid system for the photoelectrochemical immunoassay of SirT1.

Human sirtuin1 (SirT1), which is a member of the sirtuin family, plays an important role in a wide range of cellular processes. Here we demonstrate a new strategy for the photoelectrochemical assay of SirT1 in different cell lines based on a semiconductor-polymer hybrid system consisting of Au-polymer and TiO2-Au nanocomposites. Au-polymer (GC-HBAP) hybrids were synthesized from crosslinked hyperbranched azo-polymer and gold colloids and then used as an immobilization platform for SirT1 antibody. Gold-doped TiO2 (TiO2-Au) nanocomposites were prepared as the photoelectrochemical labels for signal readout in the sandwiched immunoassay. The integration of GC-HBAP with TiO2-Au facilitated the electron transfer and the photoelectrocatalytic reaction, resulting in good analytical performance with high sensitivity, selectivity and rapid response for the analysis of SirT1 levels in different cell lines. This proposed semiconductor-polymer system might open a new perspective for the development of a highly sensitive photoelectrochemical immunosensor, and have potentially promising applications in assays of other proteins.

Sensitive electrochemical immunosensor for α-synuclein based on dual signal amplification using PAMAM dendrimer-encapsulated Au and enhanced gold nanoparticle labels.

A novel electrochemical immunosensor for sensitive detection of α-synuclein (α-SYN), a very important neuronal protein, has been developed based on dual signal amplification strategy. Herein, G4-polyamidoamine dendrimer-encapsulated Au nanoparticles (PAMAM-Au nanocomposites) were covalently bound on the poly-o-aminobenzoic acid (poly-o-ABA), which was initially electropolymerized on the electrode surface to perform abundant carboxyl groups. The formed immunosensor platform, PAMAM-Au, was proved to provide numerous amino groups to allow highly dense immobilization of antigen, and facilitate the improvement of electrochemical responses as well. Subsequently, the enhanced gold nanoparticle labels ({HRP-Ab(2)-GNPs}) were fabricated by immobilizing horseradish peroxidase-secondary antibody (HRP-Ab(2)) on the surface of gold nanoparticles (GNPs). After an immunoassay process, the {HRP-Ab(2)-GNPs} labels were introduced onto the electrode surface, and produced an electrocatalytic response by reduction of hydrogen peroxide (H(2)O(2)) in the presence of enzymatically oxidized thionine. On the basis of the dual signal amplification of PAMAM-Au and {HRP-Ab(2)-GNPs} labels, the designed immunosensor displayed an excellent analytical performance with high sensitivity and stability. This developed strategy was successfully proved as a simple, cost-effective method, and could be easily extended to other protein analysis schemes.

A dual enzymatic-biosensor for simultaneous determination of glucose and cholesterol in serum and peritoneal macrophages of diabetic mice: evaluation of the diabetes-accelerated atherosclerosis risk.

In this paper, a novel dual enzymatic-biosensor is described for simultaneous determination of glucose and cholesterol in serum and peritoneal macrophages (PMs) of diabetic mice to evaluate the risk of diabetes-accelerated atherosclerosis. The biosensor was constructed by a three-step method. First, a poly-thionine (PTH) film was assembled on the surface of glassy carbon electrode by cyclic voltammetric electropolymerization of thionine, which serves as an electron transfer mediator (ETM). Second, gold nanoparticles (GNPs) were covered on the surface of PTH facilitating the electron transfer between glucose oxidase (GOx), cholesterol oxidase (ChOx) and electrode. Finally, the enzymes, GOx, cholesterol esterase (ChE), and ChOx, were covalently attached to the PTH layer through a chitosan (CH) linker. The PTH coupled with GNPs provides good selectivity, high sensitivity and little crosstalk for the dual enzymatic-biosensor. The developed biosensor had good electrocatalytic activity toward the oxidations of glucose and cholesterol, exhibiting a linear range from 0.008 mM to 6.0 mM for glucose with a detection limit of 2.0 µM, and a linear range from 0.002 mM to 1.0 mM for cholesterol with a detection limit of 0.6 µM. The results of the diabetic mice demonstrated that the cholesterol level did not change obviously with the increase of glucose level in serum, while the cholesterol level was induced with the increase of the glucose level in PMs. Previous studies have shown that the large accumulation of cholesterol in macrophage could lead to macrophage foam cell formation, which is the hallmark of early atherosclerosis. This study provides useful further evidences for the development of diabetes-accelerated atherosclerosis.