Graphene-Based Field-Effect Transistor for Ultrasensitive Immunosensing of SARS-CoV-2 Spike S1 Antigen.

Coronavirus disease (COVID-19) is an infectious disease that has posed a global health challenge caused by the SARS-CoV-2 virus. Early management and diagnosis of SARS-CoV-2 are crucial for the timely treatment, traceability, and reduction of viral spread. We have developed a rapid method using a Graphene-based Field-Effect Transistor (Gr-FET) for the ultrasensitive detection of SARS-CoV-2 Spike S1 antigen (S1-Ag). The in-house developed antispike S1 antibody (S1-Ab) was covalently immobilized on the surface of a carboxy functionalized graphene channel using carbodiimide chemistry. Ultraviolet-visible spectroscopy, Fourier-Transform Infrared Spectroscopy, X-ray Photoelectron Spectroscopy (XPS), Atomic Force Microscopy (AFM), Optical Microscopy, Raman Spectroscopy, Scanning Electron Microscopy (SEM), Enzyme-Linked Immunosorbent Assays (ELISA), and device stability studies were conducted to characterize the bioconjugation and fabrication process of Gr-FET. In addition, the electrical response of the device was evaluated by monitoring the change in resistance caused by Ag-Ab interaction in real time. For S1-Ag, our Gr-FET devices were tested in the range of 1 fM to 1 µM with a limit of detection of 10 fM in the standard buffer. The fabricated devices are highly sensitive, specific, and capable of detecting low levels of S1-Ag.

Label free detection of SARS CoV-2 Receptor Binding Domain (RBD) protein by fabrication of gold nanorods deposited on electrochemical immunosensor (GDEI).

Coronavirus Disease 2019 (COVID-19) pandemic has shown the need for early diagnosis to manage infectious disease outbreaks. Here, we report a label free electrochemical Fluorine-Doped Tin Oxide (FTO) Immunosensor coupled with gold nanorods (GNRs) as an electron carrier for ultrasensitive detection of the Receptor Binding Domain (RBD) of SARS CoV-2 Spike protein. The RBD gene was cloned, and expressed in-house with confirmed molecular weight of ∼31 kDa via Sodium Dodecyl Sulphate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and Matrix-Assisted Laser Desorption/Ionization-Time of Flight (MALDI-TOF). RBD antibodies (Ab) were generated to be used as a bioreceptor for sensor fabrication, and characterized using SDS-PAGE, Western Blot, and Enzyme-Linked Immunosorbent Assay (ELISA). GNRs were fabricated on the electrode surface, followed by immobilization of RBD Ab. The conjugation steps were confirmed by UV-Vis Spectroscopy, Dynamic Light Scattering (DLS), Atomic Force Microscopy (AFM), Transmission Electron Microscopy (TEM), Cyclic Voltammetry (CV), and Differential Pulse Voltammetry (DPV). The fabricated electrode was further optimized for maximum efficiency and output. The detection limit of the developed electrode was determined as 0.73 fM for RBD antigen (Ag). Furthermore, the patient nasopharyngeal samples were collected in Viral Transport Media (VTM), and tested on the sensor surface that resulted in detection of SARS CoV-2 within 30 s, which was further validated via Reverse Transcription-Polymerase Chain Reaction (RT-PCR). Moreover, the immunosensor showed good repeatability, storage stability, and minimal cross reactivity against Middle East Respiratory Syndrome (MERS) spike protein. Along with ease of fabrication, the electrodes show future miniaturization potential for extensive and rapid screening of populations for COVID-19.

A Recent Update on Advanced Molecular Diagnostic Techniques for COVID-19 Pandemic: An Overview.

Coronavirus disease 2019 (COVID-19), which started out as an outbreak of pneumonia, has now turned into a pandemic due to its rapid transmission. Besides developing a vaccine, rapid, accurate, and cost-effective diagnosis is essential for monitoring and combating the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its related variants on time with precision and accuracy. Currently, the gold standard for detection of SARS-CoV-2 is Reverse Transcription Polymerase Chain Reaction (RT-PCR), but it lacks accuracy, is time-consuming and cumbersome, and fails to detect multi-variant forms of the virus. Herein, we have summarized conventional diagnostic methods such as Chest-CT (Computed Tomography), RT-PCR, Loop Mediated Isothermal Amplification (LAMP), Reverse Transcription-LAMP (RT-LAMP), as well new modern diagnostics such as CRISPR-Cas-based assays, Surface Enhanced Raman Spectroscopy (SERS), Lateral Flow Assays (LFA), Graphene-Field Effect Transistor (GraFET), electrochemical sensors, immunosensors, antisense oligonucleotides (ASOs)-based assays, and microarrays for SARS-CoV-2 detection. This review will also provide an insight into an ongoing research and the possibility of developing more economical tools to tackle the COVID-19 pandemic.

Label-free detection of SARS-CoV-2 Spike S1 antigen triggered by electroactive gold nanoparticles on antibody coated fluorine-doped tin oxide (FTO) electrode.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, also known as 2019-nCov or COVID-19) outbreak has become a huge public health issue due to its rapid transmission making it a global pandemic. Here, we report fabricated fluorine doped tin oxide (FTO) electrodes/gold nanoparticles (AuNPs) complex coupled with in-house developed SARS-CoV-2 spike S1 antibody (SARS-CoV-2 Ab) to measure the response with Cyclic Voltammetry (CV) and Differential Pulse Voltammetry (DPV). The biophysical characterisation of FTO/AuNPs/SARS-CoV-2Ab was done via UV-Visible spectroscopy, Dynamic Light Scattering (DLS), and Fourier Transform Infrared Spectroscopy (FT-IR). The fabricated FTO/AuNPs/SARS-CoV-2Ab immunosensor was optimised for response time, antibody concentration, temperature, and pH. Under optimum conditions, the FTO/AuNPs/Ab based immunosensor displayed high sensitivity with limit of detection (LOD) up to 0.63 fM in standard buffer and 120 fM in spiked saliva samples for detection of SARS-CoV-2 spike S1 antigen (Ag) with negligible cross reactivity Middle East Respiratory Syndrome (MERS) spike protein. The proposed FTO/AuNPs/SARS-CoV-2Ab based biosensor proved to be stable for up to 4 weeks and can be used as an alternative non-invasive diagnostic tool for the rapid, specific and sensitive detection of SARS-CoV-2 Spike Ag traces in clinical samples.