Paper-based electrochemical biosensors for the diagnosis of viral diseases.

Paper-based electrochemical analytical devices (ePADs) have gained significant interest as promising analytical units in recent years because they can be fabricated in simple ways, are low-cost, portable, and disposable platforms that can be applied in various fields. In this sense, paper-based electrochemical biosensors are attractive analytical devices since they can promote diagnose several diseases and potentially allow decentralized analysis. Electrochemical biosensors are versatile, as the measured signal can be improved by using mainly molecular technologies and nanomaterials to attach biomolecules, resulting in an increase in their sensitivity and selectivity. Additionally, they can be implemented in microfluidic devices that drive and control the flow without external pumping and store reagents, and improve the mass transport of analytes, increasing sensor sensitivity. In this review, we focus on the recent developments in electrochemical paper-based devices for viruses' detection, including COVID-19, Dengue, Zika, Hepatitis, Ebola, AIDS, and Influenza, among others, which have caused impacts on people's health, especially in places with scarce resources. Also, we discuss the advantages and disadvantages of the main electrode's fabrication methods, device designs, and biomolecule immobilization strategies. Finally, the perspectives and challenges that need to be overcome to further advance paper-based electrochemical biosensors' applications are critically presented.

Rapid Detection of Anti-SARS-CoV-2 Antibodies with a Screen-Printed Electrode Modified with a Spike Glycoprotein Epitope.

BACKGROUND: The coronavirus disease of 2019 (COVID-19) is caused by an infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). It was recognized in late 2019 and has since spread worldwide, leading to a pandemic with unprecedented health and financial consequences. There remains an enormous demand for new diagnostic methods that can deliver fast, low-cost, and easy-to-use confirmation of a SARS-CoV-2 infection. We have developed an affordable electrochemical biosensor for the rapid detection of serological immunoglobulin G (IgG) antibody in sera against the spike protein. MATERIALS AND METHODS: A previously identified linear B-cell epitope (EP) specific to the SARS-CoV-2 spike glycoprotein and recognized by IgG in patient sera was selected for the target molecule. After synthesis, the EP was immobilized onto the surface of the working electrode of a commercially available screen-printed electrode (SPE). The capture of SARS-CoV-2-specific IgGs allowed the formation of an immunocomplex that was measured by square-wave voltammetry from its generation of hydroquinone (HQ). RESULTS: An evaluation of the performance of the EP-based biosensor presented a selectivity and specificity for COVID-19 of 93% and 100%, respectively. No cross-reaction was observed to antibodies against other diseases that included Chagas disease, Chikungunya, Leishmaniosis, and Dengue. Differentiation of infected and non-infected individuals was possible even at a high dilution factor that decreased the required sample volumes to a few microliters. CONCLUSION: The final device proved suitable for diagnosing COVID-19 by assaying actual serum samples, and the results displayed good agreement with the molecular biology diagnoses. The flexibility to conjugate other EPs to SPEs suggests that this technology could be rapidly adapted to diagnose new variants of SARS-CoV-2 or other pathogens.

A Pencil-Lead Immunosensor for the Rapid Electrochemical Measurement of Anti-Diphtheria Toxin Antibodies.

Diphtheria is a vaccine-preventable disease, yet immunization can wane over time to non-protective levels. We have developed a low-cost, miniaturized electroanalytical biosensor to quantify anti-diphtheria toxin (DTx) immunoglobulin G (anti-DTx IgG) antibody to minimize the risk for localized outbreaks. Two epitopes specific to DTx and recognized by antibodies generated post-vaccination were selected to create a bi-epitope peptide, biEP, by synthesizing the epitopes in tandem. The biEP peptide was conjugated to the surface of a pencil-lead electrode (PLE) integrated into a portable electrode holder. Captured anti-DTx IgG was measured by square wave voltammetry from the generation of hydroquinone (HQ) from the resulting immunocomplex. The performance of the biEP reagent presented high selectivity and specificity for DTx. Under the optimized working conditions, a logarithmic calibration curve showed good linearity over the concentration range of 10-5-10-1 IU mL-1 and achieved a limit of detection of 5 × 10-6 IU mL-1. The final device proved suitable for interrogating the immunity level against DTx in actual serum samples. Results showed good agreement with those obtained from a commercial enzyme-linked immunosorbent assay. In addition, the flexibility for conjugating other capture molecules to PLEs suggests that this technology could be easily adapted to the diagnoses of other pathogens.

Combined Colorimetric and Electrochemical Measurement Paper-Based Device for Chemometric Proof-of-Concept Analysis of Cocaine Samples.

Cocaine (COC) is one of the most widely consumed illegal drugs around the world. Street COC is commonly adulterated with pharmaceutical compounds that mimic or intensify the COC's sensory effect. Adulteration is performed to increase the profit of criminal organizations and each one has their own way of doing it. Therefore, determining the composition of seized COC samples (chemical profile) provides evidence for the police to track criminal organization networks and their activity patterns. Using filter paper as a substrate, we developed a multiple detection paper-based analytical device (PAD) that combines colorimetric and electrochemical measurements to discriminate COC samples according to adulterant's content. A regular graphite lead modified with a gold film made from Au leaf (graphite/Au) to improve electron transfer was used as a working electrode. Silver and Ag/AgCl were used as auxiliary and reference electrodes, respectively. The colorimetric device was patterned using a laser cutter and coupled to the electrochemical device using a double-sided tape, allowing simultaneous analysis to gather more analytical information about COC samples. Graphite/Au was characterized by scanning and transmission electron microscopies and electrochemical assays. The simultaneous colorimetric and electrochemical analyses combined to principal component analysis improved the analytical characterization of COC trial samples and provided a fast discrimination based on the assembled database.

Enhanced performance of pencil-drawn paper-based electrodes by laser-scribing treatment.

Electrochemical Paper-based Analytical Devices (ePADs) are an alternative to traditional portable analytical techniques due to features such as low-cost, easy surface modification with different materials, and high sensitivity. A fast and simple method to fabricate enhanced ePADs using pencil-drawing which involves the CO2 laser treatment of the carbon surface deposited on paper is described. The electrochemical performances of the devices were evaluated using cyclic voltammetry (CV) with different redox probes and electrochemical impedance spectroscopy (EIS). The electrochemical results show that a treated surface presents a lower resistance to charge transfer and changes the approach of the probe and the overlap of its orbitals with the electrode. To investigate the effects of the laser treatment process, chemical and structural characteristics were evaluated using scanning electron microscopy (SEM), X-ray photoelectron spectroscopy (XPS), and Raman spectroscopy. These results indicated that laser treatment promoted the restoration of carbon-carbon double bonds and removed a thin layer of nanodebris present in commercial pencils, resulting in an improvement of the electrochemical kinetics. As a proof-of-concept, the Pencil-Drawing Electrode (PDE) was used for the detection and quantification of furosemide (FUR) in a sample of synthetic urine, exhibiting a limit of detection (LOD) of 2.4 × 10-7 mol L-1. The percentages of recovery of the FUR added to the samples A and B were 95% and 110%, respectively. The analysis using CO2 laser-treated PDE resulted in a fast, simple, and reliable method for this doping agent.

Single-Step Reagentless Laser Scribing Fabrication of Electrochemical Paper-Based Analytical Devices.

A single-step laser scribing process is used to pattern nanostructured electrodes on paper-based devices. The facile and low-cost technique eliminates the need for chemical reagents or controlled conditions. This process involves the use of a CO2 laser to pyrolyze the surface of the paperboard, producing a conductive porous non-graphitizing carbon material composed of graphene sheets and composites with aluminosilicate nanoparticles. The new electrode material was extensively characterized, and it exhibits high conductivity and an enhanced active/geometric area ratio; it is thus well-suited for electrochemical purposes. As a proof-of-concept, the devices were successfully employed for different analytical applications in the clinical, pharmaceutical, food, and forensic fields. The scalable and green fabrication method associated with the features of the new material is highly promising for the development of portable electrochemical devices.