A Review from a Clinical Perspective: Recent Advances in Biosensors for the Detection of L-Amino Acids.

The field of biosensors is filled with reports and designs of various sensors, with the vast majority focusing on glucose sensing. However, in addition to glucose, there are many other important analytes that are worth investigating as well. In particular, L-amino acids appear as important diagnostic markers for a number of conditions. However, the progress in L-amino acid detection and the development of biosensors for L-amino acids are still somewhat insufficient. In recent years, the need to determine L-amino acids from clinical samples has risen. More clinical data appear to demonstrate that abnormal concentrations of L-amino acids are related to various clinical conditions such as inherited metabolic disorders, dyslipidemia, type 2 diabetes, muscle damage, etc. However, to this day, the diagnostic potential of L-amino acids is not yet fully established. Most likely, this is because of the difficulties in measuring L-amino acids, especially in human blood. In this review article, we extensively investigate the 'overlooked' L-amino acids. We review typical levels of amino acids present in human blood and broadly survey the importance of L-amino acids in most common conditions which can be monitored or diagnosed from changes in L-amino acids present in human blood. We also provide an overview of recent biosensors for L-amino acid monitoring and their advantages and disadvantages, with some other alternative methods for L-amino acid quantification, and finally we outline future perspectives related to the development of biosensing devices for L-amino acid monitoring.

Capacitance-Based Biosensor for the Measurement of Total Loss of L-Amino Acids in Human Serum during Hemodialysis.

In this paper, we present a biosensor based on a gold nanoparticle (AuNP)-modified Pt electrode with an adjusted membrane containing cross-linked L-amino acid oxidase for the detection and quantification of total L-amino acids. The designed biosensor was tested and characterized using the capacitance-based principle, capacitance measurements after electrode polarization, disconnection from the circuit, and addition of the respective amount of the analyte. The method was implemented using the capacitive and catalytic properties of the Pt/AuNP electrode; nanostructures were able to store electric charge while at the same time catalyzing the oxidation of the redox reaction intermediate H2O2. In this way, the Pt/AuNP layer was charged after the addition of analytes, allowing for much more accurate measurements for samples with low amino acid concentrations. The combined biosensor electrode with the capacitance-based measurement method resulted in high sensitivity and a low limit of detection (LOD) for hydrogen peroxide (4.15 µC/µM and 0.86 µM, respectively) and high sensitivity, a low LOD, and a wide linear range for L-amino acids (0.73 µC/µM, 5.5 µM and 25-1500 µM, respectively). The designed biosensor was applied to measure the relative loss of amino acids in patients undergoing renal replacement therapy by analyzing amino acid levels in diluted serum samples before and after entering/leaving the hemodialysis apparatus. In general, the designed biosensor in conjunction with the proposed capacitance-based method was clinically tested and could also be applied for the detection of other analytes using analyte-specific oxidases.

Biosensor prototype for rapid detection and quantification of DNase activity.

DNases are enzymes that cleave phosphodiesteric bonds of deoxyribonucleic acid molecules and are found everywhere in nature, especially in bodily fluids, i.e., saliva, blood, or sweat. Rapid and sensitive detection of DNase activity is highly important for quality control in the pharmaceutical and biotechnology industries. For clinical diagnostics, recent reports indicate that increased DNase activity could be related to various diseases, such as cancers. In this paper, we report a new bioelectronic device for the determination of nuclease activity in various fluids. The system consists of a sensor electrode, a custom design DNA target to maximize the DNase cleavage rate, a signal analysis algorithm, and supporting electronics. The developed sensor enables the determination of DNase activity in the range of 3.4 × 10-4 - 3.0 × 10-2 U mL-1 with a limit of detection of up to 3.4 × 10-4 U mL-1. The sensor was tested by measuring nuclease activity in real human saliva samples and found to demonstrate high accuracy and reproducibility compared to the industry standard DNaseAlert™ï¸. Finally, the entire detection system was implemented as a prototype device system utilizing single-use electrodes, custom-made cells, and electronics. The developed technology can improve nuclease quality control processes in the pharmaceutical/biotechnology industry and provide new insights into the importance of nucleases for medical applications.

Glucose-to-Resistor Transduction Integrated into a Radio-Frequency Antenna for Chip-less and Battery-less Wireless Sensing.

To maximize the potential of 5G infrastructure in healthcare, simple integration of biosensors with wireless tag antennas would be beneficial. This work introduces novel glucose-to-resistor transduction, which enables simple, wireless biosensor design. The biosensor was realized on a near-field communication tag antenna, where a sensing bioanode generated electrical current and electroreduced a nonconducting antenna material into an excellent conductor. For this, a part of the antenna was replaced by a Ag nanoparticle layer oxidized to high-resistance AgCl. The bioanode was based on Au nanoparticle-wired glucose dehydrogenase (GDH). The exposure of the cathode-bioanode to glucose solution resulted in GDH-catalyzed oxidation of glucose at the bioanode with a concomitant reduction of AgCl to highly conducting Ag on the cathode. The AgCl-to-Ag conversion strongly affected the impedance of the antenna circuit, allowing wireless detection of glucose. Mimicking the final application, the proposed wireless biosensor was ultimately evaluated through the measurement of glucose in whole blood, showing good agreement with the values obtained with a commercially available glucometer. This work, for the first time, demonstrates that making a part of the antenna from the AgCl layer allows achieving simple, chip-less, and battery-less wireless sensing of enzyme-catalyzed reduction reaction.

A direct electron transfer formaldehyde dehydrogenase biosensor for the determination of formaldehyde in river water.

In this work, we report the construction of a direct electron transfer (DET) biosensor based on NAD-dependent formaldehyde dehydrogenase from Pseudomonas sp. (FDH) immobilized on the gold nanoparticle-modified gold electrode. To the best of our knowledge, a DET for FDH was achieved for the first time - the oxidation of formaldehyde started at a low electrode potential of -190 mV vs. Ag/AgCl and reached a maximum current density of 1100 nA cm-2 at 200 mV vs. Ag/AgCl. Also, the designed electrode was insensitive to substrate inhibition (in comparison to the free enzyme) and operated in solutions with formaldehyde concentrations up to 10 mM. The electrode was used and characterized as a mediatorless biosensor for the detection of formaldehyde. The biosensor demonstrated a limit of detection (0.05 mM), linear range from 0.25 to 2.0 mM, the sensitivity of 178.9 nA mM cm-2, high stability and selectivity. The biosensor has been successfully tested for the determination of added formaldehyde concentration in river water samples, thus the developed electrode could be applied for a fast, inexpensive and simple measurement of formaldehyde in various media.

Real-time glucose monitoring system containing enzymatic sensor and enzymatic reference electrodes.

Every electrochemical biosensor uses two or three electrode setup, which involves sensing electrode for a specific reaction, metal/salt reference electrode (i.e., Ag/AgCl or Hg/Hg2Cl2) for the control of the potential and, is some cases, counter electrode for the compensation of the current. This setup has significant flaws related to metal/salt reference electrodes: they are bulky and difficult to miniaturize, leak electrolyte to the medium, lose the ability to define the electrochemical potential precisely in time, consequently, have to be updated or replaced. This causes problems when the biosensor cannot be easily replaced (e.g., implanted electronics). Here we present a fully enzymatic real-time glucose monitoring system capable of referencing its own electrochemical potential. Using sensing electrode composed of wired glucose dehydrogenase and enzymatic reference electrode composed of wired laccase we have created a stable and accurate electrode system, which measured fluxes in concentration of glucose in a physiological range (3-8 mM), and demonstrated performance of the designed system in undiluted human serum. In addition, our designed enzymatic reference electrode is universal and may be applied for other biosensors, thus open possibilities for the new generation of implantable devices for healthcare monitoring.

Highly sensitive amperometric biosensor based on alcohol dehydrogenase for determination of glycerol in human urine.

In this paper we report the development of a highly sensitive amperometric glycerol biosensor based on alcohol dehydrogenase from Pseudomonas putida immobilized on graphite electrode modified with carbon nanotubes and a redox mediator tetrathiafulvalene. The designed biosensor demonstrates very high sensitivity towards glycerol (29.2 ±â€¯0.9 µA mM-1 cm-2), low limit of detection (18 µM), linear range from 0.05 to 1.0 mM, high selectivity and satisfactory stability. Biosensor has been successfully used for the determination of glycerol concentration in buffer solutions as well as in the human urine samples. Received results shows a satisfactory agreement with the control measurements carried out using colorimetric commercially available glycerol determination assay kit, thus developed biosensor can be successfully applied for measurements of glycerol concentration in human urine and may be a fast, attractive and non-invasive tool for the determination of glycerol.

Preparation and characterization of iron oxide magnetic nanoparticles functionalized by nisin.

Nisin is a known bacteriocin approved as a food additive for food preservation. It exhibits a wide spectrum antimicrobial activity against Gram-positive bacteria. Iron oxide magnetic nanoparticles were synthesized and characterized by X-ray diffraction method. A main part of iron oxide nanoparticles was found to be maghemite though a small quantity of magnetite could also be present. Magnetic nanoparticles were stabilized by citric, ascorbic, gallic or glucuronic acid coating. Stable iron oxide magnetic nanoparticles were functionalized by nisin using a simple and low cost adsorption method. Nisin loading was confirmed by FT-IR spectra, thermogravimetric analysis, dynamic light scattering and atomic force microscopy methods. Nisin-loaded iron oxide magnetic nanoparticles were stable at least six weeks as judged by the measurements of zeta-potential and hydrodynamic diameter. The antimicrobial activity of nisin-loaded iron oxide magnetic nanoparticles was demonstrated toward Gram-positive bacteria. Functionalized nanoparticles could therefore find the application as antimicrobials in innovative and emerging technologies based on the magnetic field.