Merging Lateral Flow Immunoassay with Electroanalysis as a Novel Sensing Platform: Prostate Specific Antigen Detection as Case of Study.

The COVID-19 pandemic highlighted lateral flow immunoassay (LFIA) strips as the most known point-of-care (POC) devices enabling rapid and easy detection of relevant biomarkers by nonspecialists. However, these diagnostic tests are usually associated with the qualitative detection of the biomarker of interest. Alternatively, electrochemical-based diagnostics, especially known for diabetes care, enable quantitative determination of biomarkers. From an analytical point perspective, the combination of the two approaches might represent a step forward for the POC world: in fact, electrochemical transduction is attractive to be integrated into LFIA strips due to its simplicity, high sensitivity, fast signal generation, and cost effectiveness. In this work, a LFIA strip has been combined with an electrochemical transduction, yielding an electrochemical LFIA (eLFIA). As a proof-of-concept method, the detection of prostate-specific antigen has been carried out by combining a printed-electrochemical strip with the traditional LFIA tests. The electrochemical detection has been based on the measurement of Au ions produced from the dissolution of the gold nanoparticles previously captured on the test line. The analytical performances obtained at LFIA and eLFIA were compared, highlighting how the use of differential pulse voltammetry allowed for a lower detection limit (2.5-fold), respectively, 0.38 and 0.15 ng/mL, but increasing the time of analysis. Although the correlation between the two architectures confirmed the satisfactory agreement of outputs, this technical note has been thought to provide the reader a fair statement with regard to the strength and drawbacks about combining the two (apparently) competitor devices in a diagnostics field, namely, LFIA and electrochemical strips.

Design of a printed electrochemical strip towards miRNA-21 detection in urine samples: optimization of the experimental procedures for real sample application.

MicroRNAs (miRNAs) are clinical biomarkers for various human diseases, including cancer. They have been found in liquid biopsy samples, including various bodily fluids. They often play an important role in the early diagnosis and prognosis of cancer, and the development of simple and effective analytical methods would be of pivotal importance for the entire community. The determination of these targets may be affected by the different physicochemical parameters of the specimen of interest. In this work, an electrochemical detection platform for miRNA based on a screen-printed gold electrode was developed. In the present study, miRNA-21 was selected as a model sequence, due to its role in prostate, breast, colon, pancreatic, and liver cancers. A DNA sequence modified with methylene blue (MB) was covalently bound to the electrochemical strip and used to detect the selected target miRNA-21. After optimization of selected parameters in standard solutions, including the study of the effect of pH, the presence of interferent species, and NaCl salt concentration in the background, the application of square-wave voltammetry (SWV) technique allowed the detection of miRNA-21 down to a limit in the order of 2 nM. The developed device was then applied to several urine samples. In this case too, the device showed high selectivity in the presence of the complex matrix, satisfactory repeatability, and a limit of detection in the order of magnitude of nM, similarly as what observed in standard solutions.

Printed Electrochemical Strip for the Detection of miRNA-29a: A Possible Biomarker Related to Alzheimer's Disease.

The development of electrochemical strips, as extremely powerful diagnostic tools, has received much attention in the field of sensor analysis and, in particular, the detection of nucleic acids in complex matrixes is a hot topic in the electroanalytical area, especially when directed toward the development of emerging technologies, for the purpose of facilitating personal healthcare. One of the major diseases for which early diagnosis is crucial is represented by Alzheimer's disease (AD). AD is a progressive neurodegenerative disease, and it is the most common cause of dementia worldwide. In this context microRNAs (miRNAs), which are small noncoding RNAs, have recently been highlighted for their promising role as biomarkers for early diagnosis. In particular, miRNA-29 represents a class of miRNAs known to regulate pathogenesis of AD. In this work we developed an electrochemical printed strip for the detection of miRNA-29a at low levels. The architecture was characterized by the presence of gold nanoparticles (AuNPs) and an anti-miRNA-29a probe labeled with a redox mediator. The novel analytical tool has been characterized with microscale thermophoresis and electrochemical methods, and it has been optimized by selection of the most appropriate probe density to detect low target concentration. The present tool was capable to detect miRNA-29a both in standard solution and in serum, respectively, down to 0.15 and 0.2 nM. The platform highlighted good repeatability (calculated as the relative standard deviation) of ca. 10% and satisfactory selectivity in the presence of interfering species. This work has the objective to open a way for the study and possible early diagnosis of a physically and socially devastating disease such as Alzheimer's. The results demonstrate the suitability of this approach in terms of ease of use, time of production, sensitivity, and applicability.

Technical Evaluation of a Paper-Based Electrochemical Strip to Measure Nitrite Ions in the Forensic Field.

Nitrite is a compound used as a food additive for its preservative action and coloring capability, as well as an industrial agent for its antifreezing action and for preventing corrosion, and it is also used as a pharmaceutical in cyanide detoxification therapy. However, even recently, because of its high toxicity, it has been used as a murder and suicidal agent due to its affordability and ready availability. In this technical report, we describe an electrochemical paper-based device for selectively determining nitrite in complex biofluids, such as blood, cadaveric blood, vitreous humor, serum, plasma, and urine. The approach was validated in terms of the linearity of response, selectivity, and sensitivity, and the accuracy of the determination was verified by comparing the results with a chromatographic instrumental method. A linear response was observed in the micromolar range; the sensitivity of the method expressed as the limit of detection was 0.4 µM in buffer measurements. The simplicity of use, the portability of the device, and the performance shown make the approach suitable for detecting nitrite in complex biofluids, including contexts of forensic interest, such as murders or suicides in which nitrite is used as a toxic agent. Limits of detection of ca. 1, 2, 4, 5, 3, and 4 µM were obtained in vitreous humor, urine, serum and plasma, blood, and cadaveric blood, also highlighting a satisfactory accuracy comprised between 91 and 112%.

Sustainable Copper Electrochemical Stripping onto a Paper-Based Substrate for Clinical Application.

The electroanalytical field has exploited great advantages in using paper-based substrates, even if the word "paper" might be general. In fact, the mainly adopted paper-based substrates are often chromatographic and office ones. They are characterized by the following main features (and drawbacks): chromatographic paper is well-established for storing reagents/treating samples, but the sensitivity compared to traditional screen-printed ones is lower (due to porosity), while office paper represents a sustainable alternative to plastic (with similar sensitivity), but its porosity is not enough to load reagents. To overcome the limitations that might arise due to the adoption of a type of individual paper-based substrate, herein, we describe for the first time the development of a two-dimensional merged paper-based device for electrochemical copper ion detection in serum. In this work, we report a novel configuration to produce an integrated all-in-one electrochemical device, in which no additional working medium has to be added by the end user and the sensitivity can be tuned by rapid preconcentration on porous paper, with the advantage of making the platform adaptable to real matrix scenarios. The novel architecture has been obtained by combining office paper to screen-print a sustainable and robust electrochemical strip and a chromatographic disk to (1) store the reagents, (2) collect real samples, and (3) preconcentrate the analyte of interest. The novel sensing platform has allowed us to obtain a detection limit for copper ions down to 4 ppb in all the solutions that have been investigated, namely, standard solutions and serum, and a repeatability of ca. 10% has been obtained. Inductively coupled plasma-mass spectrometry measurements confirmed the satisfactory correlation.

Electrochemical Biosensors for Tracing Cyanotoxins in Food and Environmental Matrices.

The adoption of electrochemical principles to realize on-field analytical tools for detecting pollutants represents a great possibility for food safety and environmental applications. With respect to the existing transduction mechanisms, i.e., colorimetric, fluorescence, piezoelectric etc., electrochemical mechanisms offer the tremendous advantage of being easily miniaturized, connected with low cost (commercially available) readers and unaffected by the color/turbidity of real matrices. In particular, their versatility represents a powerful approach for detecting traces of emerging pollutants such as cyanotoxins. The combination of electrochemical platforms with nanomaterials, synthetic receptors and microfabrication makes electroanalysis a strong starting point towards decentralized monitoring of toxins in diverse matrices. This review gives an overview of the electrochemical biosensors that have been developed to detect four common cyanotoxins, namely microcystin-LR, anatoxin-a, saxitoxin and cylindrospermopsin. The manuscript provides the readers a quick guide to understand the main electrochemical platforms that have been realized so far, and the presence of a comprehensive table provides a perspective at a glance.