Portable microfluidic impedance biosensor for SARS-CoV-2 detection.

Pandemics as the one we are currently facing, where fast-spreading viruses present a threat to humanity, call for simple and reliable methods to perform early diagnosis, enabling detection of very low pathogen loads even before symptoms start showing in the host. So far, standard polymerase chain reaction (PCR) is the most reliable method for doing so, but it is rather slow and needs specialized reagents and trained personnel to operate it. Additionally, it is expensive and not easily accessible. Therefore, developing miniaturized and portable sensors which perform early detection of pathogens with high reliability is necessary to not only prevent the spreading of the disease but also to monitor the effectiveness of the developed vaccines and the appearance of new pathogenic variants. Thus, in this work we develop a sensitive microfluidic impedance biosensor for the direct detection of SARS-CoV-2, towards a mobile point-of-care (POC) platform. The operational parameters are optimized with the aid of design-of-experiment (DoE), for an accurate detection of the viral antigens using electrochemical impedance spectroscopy (EIS). We perform the biodetection of buffer samples spiked with fM concentration levels and validate the biosensor in a clinical context of relevance by analyzing 15 real patient samples up to a Ct value (cycle threshold) of 27. Finally, we demonstrate the versatility of the developed platform using different settings, including a small portable potentiostat, using multiple channels for self-validation, as well as with single biosensors for a smartphone-based readout. This work contributes to the rapid and reliable diagnostics of COVID-19 and can be extended to other infectious diseases, allowing the monitoring of viral load in vaccinated and unvaccinated people to anticipate a potential relapse of the disease.

Remote Floating-Gate Field-Effect Transistor with 2-Dimensional Reduced Graphene Oxide Sensing Layer for Reliable Detection of SARS-CoV-2 Spike Proteins.

Despite intensive research of nanomaterials-based field-effect transistors (FETs) as a rapid diagnostic tool, it remains to be seen for FET sensors to be used for clinical applications due to a lack of stability, reliability, reproducibility, and scalability for mass production. Herein, we propose a remote floating-gate (RFG) FET configuration to eliminate device-to-device variations of two-dimensional reduced graphene oxide (rGO) sensing surfaces and most of the instability at the solution interface. Also, critical mechanistic factors behind the electrochemical instability of rGO such as severe drift and hysteresis were identified through extensive studies on rGO-solution interfaces varied by rGO thickness, coverage, and reduction temperature. rGO surfaces in our RFGFET structure displayed a Nernstian response of 54 mV/pH (from pH 2 to 11) with a 90% yield (9 samples out of total 10), coefficient of variation (CV) < 3%, and a low drift rate of 2%, all of which were calculated from the absolute measurement values. As proof-of-concept, we demonstrated highly reliable, reproducible, and label-free detection of spike proteins of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a saliva-relevant media with concentrations ranging from 500 fg/mL to 5 µg/mL, with an R2 value of 0.984 and CV < 3%, and a guaranteed limit of detection at a few pg/mL. Taken together, this new platform may have an immense effect on positioning FET bioelectronics in a clinical setting for detecting SARS-CoV-2.

ACE2-based capacitance sensor for rapid native SARS-CoV-2 detection in biological fluids and its correlation with real-time PCR.

The spread of the SARS-CoV-2 and its increasing threat to human health worldwide have necessitated the development of new technological tools to combat the virus. Particular emphasis is given to the development of diagnostic methods that monitor the spread of the virus rapidly and effectively. In this study, we report the development and testing of an antibody-free biosensor, based on the immobilization of ACE2 protein on the surface of gold interdigitated electrode. When the sensor was used in laboratory conditions for targeting the virus' structural spike protein, it showed a limit of detection [LOD] of 750 pg/µL/mm2. Thereafter, the response of the sensor to swab and saliva samples from hospitalized patients was examined. The virus presence in the samples was confirmed by electrical effective capacitance measurements executed on the biosensor, and correlated with real-time PCR results. We verified that the biosensor can distinguish samples that are positive for the virus from those that are negative in a total of 7 positive and 16 negative samples. In addition, the biosensor can be used for semi-quantitative measurement, since its measurements are divided into 3 areas, the negative samples, the weakly positive and the positive samples. Reproducibility of the experiments was demonstrated with at least 3 replicates and stability was tested by keeping the sensor standby for 7 days at 4 °C before repeating the experiment. This work presents a biosensor that can be used as a fast-screening test at point of care detection of SARS-CoV-2 since it needs less than 2 min to provide results and is of simple operation.