Magnetic particles (MPs) have been widely used in biological applications in recent years as a carrier for various molecules. Their big advantage is in repeated use of immobilized molecules including enzymes. Acetylcholinesterase (AChE) is an enzyme playing crucial role in neurotransmission and the enzyme is targeted by various molecules like Alzheimer's drugs, pesticides and warfare agents. In this work, an electrochemical biosensor having AChE immobilized onto MPs and stabilized through glutaraldehyde (GA) molecule was proposed for assay of the neurotoxic compounds. The prepared nanoparticles were modified by pure AChE and they were used for the measurement anti-Alzheimer's drug galantamine and carbamate pesticide carbofuran with limit of detection 1.5 µM and 20 nM, respectively. All measurements were carried out using screen-printed sensor with carbon working, silver reference, and carbon auxiliary electrode. Standard Ellman's assay was used for validation measurement of both inhibitors. Part of this work was the elimination of reversible inhibitors represented by galantamine from the active site of AChE. For this purpose, we used a lower pH to get the original activity of AChE after inhibition by galantamine. We also observed decarbamylation of the AChE-carbofuran adduct. Influence of organic solvents to AChE as well as repeatability of measurement with MPs with AChE was also established.

• Klíčová slova
• acetylcholinesterase, carbofuran, electrochemistry, galantamine, magnetic particles, nanomaterial, nanoparticles, screen-printed sensor,
• MeSH
• acetylcholinesterasa MeSH
• biosenzitivní techniky MeSH
• cholinesterasové inhibitory MeSH
• enzymy imobilizované MeSH
• nanočástice * MeSH
• organofosforové sloučeniny MeSH
• pesticidy MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetylcholinesterasa MeSH
• cholinesterasové inhibitory MeSH
• enzymy imobilizované MeSH
• organofosforové sloučeniny MeSH
• pesticidy MeSH

Smartphones are widely spread and their usage does not require any trained personnel. Recently, smartphones were successfully used in analytical chemistry as a simple detection tool in some applications. This paper focuses on immobilization of acetylcholinesterase (AChE) onto commercially available pH strips with stabilization in the gelatin membrane. AChE degrades acetylcholine into choline and acetic acid which causes color change of acid-base indicator. Smartphone served as a tool for measurement of indicator color change from red to orange while inhibitors blocked this process. AChE inhibitors were measured with limits of detection, 149 nM and 22.3 nM for galanthamine and donepezil, respectively. Organic solvents were measured for method interferences. Measurement procedure was performed on 3D printed holder and digital photography was evaluated using red-green-blue (RGB) channels. The invented assay was validated to the standard Ellman's test and verified on murine plasma samples spiked with inhibitors. We consider that the assay is fully suitable for practical performance.

• Publikační typ
• časopisecké články MeSH

The use of a cell phone as a detection system is easy, simple and does not require trained personnel, which is in contrast to standard laboratory instruments. This paper deals with immobilization of acetylcholinesterase (AChE) in a gelatin matrix, and phenol red, as an indicator of AChE activity, is used in order to establish a method that is easily compatible with a camera device. AChE splits acetylcholine into choline and acetic acid, which changes the pH of a medium, resulting in a phenol red color change. The coloration changed in presence of an AChE inhibitor. Measurements were performed on 3D-printed, tube-shaped holder, and digital photography, with subsequent analysis of red-green-blue (RGB), served for assay purposes. Calibration of AChE inhibitors, tacrine and galantamine, was performed, with limit of detection equal to 1.1 nM and 1.28 µM, respectively. Interferences were also measured, resulting in a proof-of-method stability. The method was further successfully validated for the standard Ellman's assay, and verified on murine plasma samples spiked with inhibitors.

• Klíčová slova
• acetylcholinesterase, biosensor, colorimetry, drop assay, inhibitor, phenol red, smart phone,
• Publikační typ
• časopisecké články MeSH