A Surface Plasmon Resonance Imaging (SPRI) sensor has been developed for highly selective determination of cathepsin D (Cat D) or/and E (Cat E). The sensor contains immobilised pepstatin A, which binds aspartyl proteases from solution. Pepstatin A activated with N-Hydroxysuccinimide (NHS) and N-Ethyl-N'-(3-dimethylaminopropyl) carbodiimide (EDC) was immobilized on an amine-modified gold surface. Cysteamine was used for modification of the gold surface. Pepstatin A concentration and pH of interaction were optimised. A concentration of pepstatin equal to 0.5 μg mL-1 and a pH of 3.75 were selected as optimal.The sensor's dynamic response range is between 0.25 and 1.0 ng mL-1, and the detection limit is 0.12 ng mL-1. However, the sensor cannot distinguish between Cat D and Cat E. In order to demonstrate the sensor's potential, Cat E was determined in human red blood cells, Cat D in human saliva, as well as total concentration of Cat D and Cat E in human nasal polyps.
Keywords: Biosensor, cathepsin D, cathepsin E, pepstatin A, Surface Plasmon Resonance Imaging
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