AbstractAn efficient biosensor was created for the ratiometric monitoring of Cu+ and pH in the brain using both current and potential outputs. A series of N,N-bis(2-[2-(ethylthio)ethyl])-based (NS4s) derivatives was designed for the specific recognition of Cu+. After systematically evaluating the electrochemical parameters of Cu+ oxidation by tuning alkyl chain length, polyaromatic structure, and substitute group site of NS4, N,N-bis(2-[2-(ethylthio)ethyl])-2-naphthamide (NS4-C1) was finally optimized for Cu+ detection as it showed the most negative potential and the largest current density. At the same time, 9,10-anthraquinone was used as a selective pH sensor with 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) as an internal reference. This single biosensor with both current and potential signal outputs can simultaneously determine Cu+ concentrations from 0.5 to 9.5 μm and pH values ranging from 6.0 to 8.0. The efficient biosensor was applied to the simultaneous detection of Cu+ and pH in the live brain. The average levels of Cu+ were reported for the first time in the cortex, hippocampus, and striatum in a mouse model of Alzheimer's disease.

Developing a sensitive and accurate method for Furin activity is still the bottleneck for understanding the role played by Furin in cell-surface systems and even in Alzheimer's disease. In this work, a ratiometric electrochemical biosensor was developed for sensitive and accurate determination of Furin activity in the cell based on dual signal amplification stemming from a peptide with multiple response sites and the antifouling gold nano-bellflowers (GBFs). A new peptide, HS-CMRVRR↓YKDFDFG (P3), was designed for the first time to be selectively cleaved by Furin at site↓. More importantly, this peptide P3 constitutes three amino acid residues with the –COOH group subsequently used to bind with the response molecule of ferrocene, and can remarkably improve the determination sensitivity by about 2.3 fold. Meanwhile, GBFs stabilized by PEG were taken as a second element to magnify the signal of the ferrocene group via a large ratio surface area and good conductivity, as well as an antibiofouling nanosurface to reduce the biofouling of the electrode surface in cells. This double amplification strategy can greatly enhance the sensitivity of Furin detection by 6.5-fold, which is favorable for detection of low amounts of Furin. In addition, 5′-MB-GGCGCGA(T)13-SH-3′ was co-assembled as an inner reference to provide a built-in element to correct the determination error resulting from a complicated analysis environment. Finally, this sensitive and accurate Furin biosensor was successfully applied to detect Furin activity in Furin overexpressed U251 and MDA-MB-468 cells. As far as we know, this is the first report to mention an electrochemical strategy to detect Furin activity in cells.

AbstractAn efficient biosensor was created for the ratiometric monitoring of Cu+ and pH in the brain using both current and potential outputs. A series of N,N-bis(2-[2-(ethylthio)ethyl])-based (NS4s) derivatives was designed for the specific recognition of Cu+. After systematically evaluating the electrochemical parameters of Cu+ oxidation by tuning alkyl chain length, polyaromatic structure, and substitute group site of NS4, N,N-bis(2-[2-(ethylthio)ethyl])-2-naphthamide (NS4-C1) was finally optimized for Cu+ detection as it showed the most negative potential and the largest current density. At the same time, 9,10-anthraquinone was used as a selective pH sensor with 2,2′-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) as an internal reference. This single biosensor with both current and potential signal outputs can simultaneously determine Cu+ concentrations from 0.5 to 9.5 μm and pH values ranging from 6.0 to 8.0. The efficient biosensor was applied to the simultaneous detection of Cu+ and pH in the live brain. The average levels of Cu+ were reported for the first time in the cortex, hippocampus, and striatum in a mouse model of Alzheimer's disease.

To develop in vivo monitoring meter for pH measurements is still the bottleneck for understanding the role of pH plays in the brain diseases. In this work, a selective and sensitive electrochemical pH meter was developed for real-time ratiometric monitoring of pH in different regions of rat brains upon ischemia. First, 1,2-naphthoquinone (1,2-NQ) was employed and optimized as a selective pH recognition element to establish a 2H+/2e– approach over a wide range of pH from 5.8 to 8.0. The pH meter demonstrated remarkable selectivity toward pH detection against metal ions, amino acids, reactive oxygen species, and other biological species in the brain. Meanwhile, an inner reference, 6-(ferrocenyl)hexanethiol (FcHT), was selected as a built-in correction to avoid the environmental effect through coimmobilization with 1,2-NQ. In addition, three-dimensional gold nanoleaves were electrodeposited onto the electrode surface to amplify the signal by ∼4.0-fold and the measurement was achieved down to 0.07 pH. Finally, combined with the microelectrode technique, the microelectrochemical pH meter was directly implanted into brain regions including the striatum, hippocampus, and cortex and successfully applied in real-time monitoring of pH values in these regions of brain followed by global cerebral ischemia. The results demonstrated that pH values were estimated to 7.21 ± 0.05, 7.13 ± 0.09, and 7.27 ± 0.06 in the striatum, hippocampus, and cortex in the rat brains, respectively, in normal conditions. However, pH decreased to 6.75 ± 0.07 and 6.52 ± 0.03 in the striatum and hippocampus, upon global cerebral ischemia, while a negligible pH change was obtained in the cortex.

Selective and reliable method is of great importance for in vivo analysis in the complicated brain. In this work, we developed a new approach for detection of ascorbic acid (AA) based on a new nitrogen-doped nanotube fiber (NCNF) microelectrode. After electrochemical pre-treatment, the NCNF microelectrode (e-NCNF) demonstrated high electrocatalytic capability for AA oxidation with a low working potential, 0 V, resulting in high selectivity for detection of AA level against other biological species in brain, as well as producing high sensitivity. Moreover, the e-NCNF was directly assembled by carbon nanotube bundles, free from the post-modification, thus showing high reproducibility, in which the deviation of anodic current intensity of six electrodes prepared with same method did not exceed 4% (RSD, n = 6). Based on the remarkable analytical performance, the e-NCNF with good biocompatibility was successfully applied to determine the AA level as 134 ± 7 mM, in rat brain microdialysates.