A Study on the Combination of Enzyme Stabilizers and Low Temperatures in the Long-Term Storage of Glutamate Biosensor

Abstract

The importance of physiological glutamate has been widely demonstrated in cognitive and memory processes, as well as in neurotransmission. The involvement of physiological glutamate in several pathologies has also been established. Therefore, analytical devices for studying variations in physiological glutamate are of fundamental importance, particularly in preclinical studies. The necessary knowledge to develop and characterize biosensors for glutamate detection is often restricted to only a few research groups. However, many more groups have sought to implant such analytical devices to study the glutamatergic system in vivo. On this basis, a series of studies was undertaken to explore the medium-term storage of biosensors, thereby allowing their usage results to be differentiated from their construction and characterization processes to facilitate the wider diffusion and use of such sensors. Therefore, it has become vital to determine the best storage conditions to extend the life and functionality of these biosensors, especially due to the diachronic instability of the enzyme present on the surface. In the present study, we analyzed the impact of glycols, such as glycerol and triethylene glycol, as enzyme stabilizers coupled with long-term storage at low temperatures (−20 and −80 °C) on biosensor performance. The biosensors were observed for 5 months and evaluated for their enzymatic activity by measuring the VMAX(app) and KM(app). The analytical features were also evaluated in terms of the Linear Region Slope, which is one the most important parameters for indicating the efficiency and the sensitivity of biosensors. Interestingly, both glycols proved to be capable of increasing enzymatic activity and maintaining good biosensor efficiency over time. Moreover, the combination with low-temperature storage highlighted the different behaviors of the two glycols. In particular, glycerol was more effective in stabilizing the enzyme and maintaining analytical performance when the biosensors were stored at −20 °C. Instead, triethylene glycol performed the same function as glycerol but when the biosensors were stored at −80 °C.

Sustainable Electropolymerization of Zingerone and Its C2 Symmetric Dimer for Amperometric Biosensor Films

“…Based on the particular characteristics highlighted on both polyZING and polyZING DIM ( Figure 10 ), related to good AA shielding and increased HP monitoring, glutamate biosensors were built using the polymers mentioned above and compared with the previously described PPD-based glutamate biosensors [ 13 , 14 , 53 ]: actually, PPD is the most commonly used polymer for amperometric biosensors because of its peculiarities [ 2 , 20 ].…”

“…As previously reported [ 13 , 14 , 53 ], PPD-based biosensors (n = 4) were constructed. In brief, at day 0, a PPD polymer was obtained on a 1 mm cylinder of Pt/Ir, from a 330 mM solution of OPD, previously degassed for 15 min with pure N 2 , and by applying +0.7 V for 30 min.…”

“…Thus, two layers of PEI (1%) were deposited, and then 5 layers of GluOx were stratified, with 5 min intervals between each layer. Finally, the biosensor was rapidly immersed in a TEG solution (0.1%) and let dry for 30 min, obtaining the following design [ 14 ]: Pt c /PPD/PEI (1%) 2 /GlutOx 5 /TEG (0.1%) …”

“…This biosensor exploits the capability of glutamate oxidase (GlutOx) to recognize L-glutamate in a complex matrix specifically. As previously reported [13][14][15], the quantification of L-glutamate occurs through the oxidation of HP produced during the enzymatic reaction, as follows:…”

Modern analytical and bioanalytical technologies and concepts for smart and precision farming

“…62 To extend the storage stability of enzymes and immunosensors, biosensors must be kept with a stabilizer and buffer at a low temperature. 63,64 Also, soil and water quality assessments usually cover a few sampling spots to prevent bias in the measurement results. Hence, a reusable biosensor with high repeatability would be appreciated for multiple measurements at specific spots to reduce cost.…”