A highly sensitive and simply operated protease sensor toward point-of-care testing
| Autor: | Ji-Joon Song, Seonhwa Park, Jeongwook Seo, Haesik Yang, Yu Mi Shin |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
Proteases
Botulinum Toxins medicine.medical_treatment Biosensing Techniques 02 engineering and technology Aminophenols 010402 general chemistry 01 natural sciences Biochemistry Dithiothreitol Analytical Chemistry chemistry.chemical_compound Adsorption Limit of Detection Endopeptidases Exopeptidases Electrochemistry medicine Environmental Chemistry Amino Acid Sequence Electrodes Spectroscopy Detection limit Protease Chromatography biology Chemistry Tin Compounds Exopeptidase 021001 nanoscience & nanotechnology Endopeptidase 0104 chemical sciences Point-of-Care Testing Proteolysis biology.protein 0210 nano-technology Avidin |
| Zdroj: | The Analyst. 141:2481-2486 |
| ISSN: | 1364-5528 0003-2654 |
| DOI: | 10.1039/c6an00251j |
| Popis: | Protease sensors for point-of-care testing (POCT) require simple operation, a detection period of less than 20 minutes, and a detection limit of less than 1 ng mL(-1). However, it is difficult to meet these requirements with protease sensors that are based on proteolytic cleavage. This paper reports a highly reproducible protease sensor that allows the sensitive and simple electrochemical detection of the botulinum neurotoxin type E light chain (BoNT/E-LC), which is obtained using (i) low nonspecific adsorption, (ii) high signal-to-background ratio, and (iii) one-step solution treatment. The BoNT/E-LC detection is based on two-step proteolytic cleavage using BoNT/E-LC (endopeptidase) and l-leucine-aminopeptidase (LAP, exopeptidase). Indium-tin oxide (ITO) electrodes are modified partially with reduced graphene oxide (rGO) to increase their electrocatalytic activities. Avidin is then adsorbed on the electrodes to minimize the nonspecific adsorption of proteases. Low nonspecific adsorption allows a highly reproducible sensor response. Electrochemical-chemical (EC) redox cycling involving p-aminophenol (AP) and dithiothreitol (DTT) is performed to obtain a high signal-to-background ratio. After adding a C-terminally AP-labeled oligopeptide, DTT, and LAP simultaneously to a sample solution, no further treatment of the solution is necessary during detection. The detection limits of BoNT/E-LC in phosphate-buffered saline are 0.1 ng mL(-1) for an incubation period of 15 min and 5 fg mL(-1) for an incubation period of 4 h. The detection limit in commercial bottled water is 1 ng mL(-1) for an incubation period of 15 min. The developed sensor is selective to BoNT/E-LC among the four types of BoNTs tested. These results indicate that the protease sensor meets the requirements for POCT. |
| Databáze: | OpenAIRE |
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