Designing and fabrication of electrochemical nano-biosensor for the fast detection of SARS-CoV-2-RNA.

SARS-CoV-2 caused a global panic among populations. Rapid diagnostic procedures for the virus are crucial for disease control. Thus, the designed signature probe from a highly conserved region of the virus was chemically immobilized onto the nanostructured-AuNPs/WO3-screen printed electrodes. Different concentrations of the matched oligonucleotides were spiked to test the specificity of the hybridization affinity whereas the electrochemical impedance spectroscopy was used for tracking the electrochemical performance. After a full assay optimization, limits of detection and quantification were calculated based on linear regression and were valued at 298 and 994 fM, respectively. Further, the high performance of the fabricated RNA-sensor chips was confirmed after testing the interference status in the presence of the mismatched oligos in one nucleotide and completely one. Worthy to mention that the single-stranded matched oligos can be hybridized to the immobilized probe in 5 min at room temperature. The designed disposable sensor chips are capable of detecting the virus genome directly. Therefore, the chips are a rapid tool for SARS-CoV-2 detection.

How an ACE2 mimicking epitope-MIP nanofilm recognizes template-related peptides and the receptor binding domain of SARS-CoV-2.

Here we aim to gain a mechanistic understanding of the formation of epitope-imprinted polymer nanofilms using a non-terminal peptide sequence, i.e. the peptide GFNCYFP (G485 to P491) of the SARS-CoV-2 receptor binding domain (RBD). This epitope is chemisorbed on the gold surface through the central cysteine 488 followed by the electrosynthesis of a ∼5 nm thick polyscopoletin film around the surface confined templates. The interaction of peptides and the parent RBD and spike protein with the imprinted polyscopoletin nanofilm was followed by electrochemical redox marker gating, surface enhanced infrared absorption spectroscopy and conductive AFM. Because the use of non-terminal epitopes is especially intricate, here we characterize the binding pockets through their interaction with 5 peptides rationally derived from the template sequence, i.e. implementing central single amino acid mismatch as well as elongations and truncations at its C- and N- termini. Already a single amino acid mismatch, i.e. the central Cys488 substituted by a serine, results in ca. 15-fold lower affinity. Further truncation of the peptides to tetrapeptide (EGFN) and hexapeptide (YFPLQS) results also in a significantly lower affinity. We concluded that the affinity towards the different peptides is mainly determined by the four amino acid motif CYFP present in the sequence of the template peptide. A higher affinity than that for the peptides is found for the parent proteins RBD and spike protein, which seems to be due to out of cavity effects caused by their larger footprint on the nanofilm surface.

Advances in Electrochemical Nano-Biosensors for Biomedical and Environmental Applications: From Current Work to Future Perspectives.

Modern life quality is strongly supported by the advances made in biosensors, which has been attributed to their crucial and viable contribution in point-of-care (POC) technology developments. POC devices are exploited for the fast tracing of disease progression, rapid analysis of water, and food quality assessment. Blood glucose meters, home pregnancy strips, and COVID-19 rapid tests all represent common examples of successful biosensors. Biosensors can provide great specificity due to the incorporation of selective bio-recognition elements and portability at significantly reduced costs. Electrochemical biosensor platforms are one of the most advantageous of these platforms because they offer many merits, such as being cheap, selective, specific, rapid, and portable. Furthermore, they can be incorporated into smartphones and various analytical approaches in order to increase their sensitivity and many other properties. As a very broad and interdisciplinary area of research and development, biosensors include all disciplines and backgrounds from materials science, chemistry, physics, medicine, microbiology/biology, and engineering. Accordingly, in this state-of-the-art article, historical background alongside the long journey of biosensing construction and development, starting from the Clark oxygen electrode until reaching highly advanced wearable stretchable biosensing devices, are discussed. Consequently, selected examples among the miscellaneous applications of nanobiosensors (such as microbial detection, cancer diagnosis, toxicity analysis, food quality-control assurance, point of care, and health prognosis) are described. Eventually, future perspectives for intelligent biosensor commercialization and exploitation in real-life that is going to be supported by machine learning and artificial intelligence (AI) are stated.

Disposable impedimetric nano-immunochips for the early and rapid diagnosis of Vitamin-D deficiency

Deficiency of Vitamin D is often misdiagnosed as its symptoms may not be manifested for several years. Long-standing of this deficiency is associated with cancer, osteoporosis, diabetes, blood clotting disorders, and recently it was involved in the progression of coronavirus 2019 infection. Therefore, the development of a point-of-care test for Vitamin D is extremely necessary to achieve early, faster, and personal diagnosis. Serum 25-hydroxyvitamin-D3 (25(OH)D3) is clinically considered as one of the remarkable biomarkers for the direct determination of vitamin D level.Herein, nanostructured-immunosensors were designed for the label-free impedimetric determination of 25(OH)D biomarker in human serum. To fabricate the sensing probe, the targeting antibody (anti-25(OH)D3) was chemically conjugated on the nanocomposite surface (AuNPs/RGO-SeO2) using 4-aminothiophenol (4-ATP) and glutaraldehyde (GA) as cross-linking agents. Through the assay optimization, screening of nanomaterials, determining the best ratio of the nanocomposite, adjusting the time of SAM-formation, and identifying the degree of cross-reactivity with other non-targeting molecules were studied and optimized. Eventually, a linear dynamic range of 0.05–200 ng/mL, and the lower limit of detection of 0.01 ng/mL was achieved. Furthermore, the proposed immunosensor was tested for the detection of 25(OH)D3 in human serum samples, and the results were in a good agreement with the result obtained by the ELISA test. Providing high selectivity, reproducibility, and stability, this designed sensor is recommended for clinical diagnosis of Vitamin D.

SARS-CoV-2-Impedimetric Biosensor: Virus-Imprinted Chips for Early and Rapid Diagnosis.

Due to the current global SARS-CoV-2 pandemic, rapid and accurate diagnostic tools are needed to prevent the spread of COVID-19 across the globe. An electrochemical sensing platform was constructed using CNTs/WO3-screen printed electrodes for imprinting the complete virus particles (SARS-CoV-2 particles) within the polymeric matrix to create viral complementary binding sites. The sensor provided high selectivity toward the target virus over other tested human corona and influenza respiratory interference viruses. The sensitivity performance of the sensor chips was evaluated using different viral concentrations, while the limits of detection and quantification were 57 and 175 pg/mL, respectively. Reaching this satisfied low detection limit (almost 27-fold more sensitive than the RT-PCR), the sensor was applied in clinical specimens obtained from SARS-CoV-2 suspected cases. Thus, dealing directly with clinical samples on the chip could be provided as a portable device for instantaneous and simple point of care in hospitals, airports, and hotspots.

Point-of-Care Diagnostics of COVID-19: From Current Work to Future Perspectives.

Coronaviruses have received global concern since 2003, when an outbreak caused by SARS-CoV emerged in China. Later on, in 2012, the Middle-East respiratory syndrome spread in Saudi Arabia, caused by MERS-CoV. Currently, the global crisis is caused by the pandemic SARS-CoV-2, which belongs to the same lineage of SARS-CoV. In response to the urgent need of diagnostic tools, several lab-based and biosensing techniques have been proposed so far. Five main areas have been individuated and discussed in terms of their strengths and weaknesses. The cell-culture detection and the microneutralization tests are still considered highly reliable methods. The genetic screening, featuring the well-established Real-time polymerase chain reaction (RT-PCR), represents the gold standard for virus detection in nasopharyngeal swabs. On the other side, immunoassays were developed, either by screening/antigen recognition of IgM/IgG or by detecting the whole virus, in blood and sera. Next, proteomic mass-spectrometry (MS)-based methodologies have also been proposed for the analysis of swab samples. Finally, virus-biosensing devices were efficiently designed. Both electrochemical immunosensors and eye-based technologies have been described, showing detection times lower than 10 min after swab introduction. Alternative to swab-based techniques, lateral flow point-of-care immunoassays are already commercially available for the analysis of blood samples. Such biosensing devices hold the advantage of being portable for on-site testing in hospitals, airports, and hotspots, virtually without any sample treatment or complicated lab precautions.