Trypsin is the most important digestive enzyme produced in the pancreas, and is a useful biomarker for pancreatitis. In this work, a rapid and sensitive method for the quantitative determination of trypsin activity is developed by using a biological alpha-hemolysin protein nanopore. Due to its much larger molecular diameter than the narrow pore constriction, trypsin itself cannot transport through the alpha-hemolysin channel. Hence, an indirect trypsin detection method is developed by monitoring its proteolytic cleavage of a lysine-containing peptide substrate. Based on the current modulations produced by the translocation of the substrate degradation products in the nanopore, the activity levels of trypsin could be determined. The method is rapid and highly sensitive, with picomolar concentrations of trypsin detected in minutes. In addition, the effects of cation and temperature on the sensor sensitivity, trypsin inhibition, and serum sample analysis are also investigated.

A label-free method for the measurement of the activity of HIV-1 protease is developed by real-time monitoring of the cleavage of a peptide substrate by HIV-1 protease in a nanopore. The method is rapid and sensitive: picomolar concentrations of HIV-1 protease could be detected in ~10 minutes. Simulated clinical samples are analyzed, and the activity of HIV-1 protease could be accurately detected. Our developed nanopore sensor design strategy should find useful application in the development of stochastic sensors for other proteases of medical, pharmaceutical, and biological importance.

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Scheme 1. Schematic illustration of a single nanopore sensor (left), typical single-channel recording trace segments (middle), and the constructed 3D plots of event counts versus residence time versus blockage amplitude for metal ion detection and differentiation (right). The ionic current through the nanopore is maintained by applying a voltage bias between two Ag/AgCl electrodes in an electrolyte solution.

To assess the land use effects on soil micronutrients, this study examined the profile variation and storage of DTPAextractable iron, manganese, copper and zinc at the depth of 0-150 cm of an aquic brown soil under four land use patterns, i.e. paddy, maize, and fallow fields and woodland, over 14 years in an ecological experimental station of northeastern China. Results showed that land use effect, soil depth, and their interactions on micronutrients were significantly different, and they were decreased with soil depth. Micronutrient storages in woodland and fallow field were significantly greater than in paddy field (P < 0.05), and significantly or comparatively greater than in maize field. Micronutrients were positively correlated with soil organic carbon, but negatively with soil pH in profiles. Plant cycling and soil pH may contribute a lot in enhancing soil micronutrient levels in woodland and fallow field, while the lower organic matter content and higher soil pH may inhibit the micronutrient availability in paddy field. The study suggested that the profile distribution of soil micronutrients was mainly controlled by biological cycling, anthropogenic disturbance and leaching and strongly affected by land uses.

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