key: cord-0776295-6ad2ax8n authors: Roberts, Akanksha; Mahari, Subhasis; Shahdeo, Deepshikha; Gandhi, Sonu title: Label-free detection of SARS-CoV-2 Spike S1 Antigen triggered by electroactive gold nanoparticles on Antibody coated Fluorine-Doped Tin Oxide (FTO) Electrode date: 2021-10-21 journal: Anal Chim Acta DOI: 10.1016/j.aca.2021.339207 sha: 200570a001c6bee3e4cb9e09117c5a972c027e13 doc_id: 776295 cord_uid: 6ad2ax8n 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 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. CoV-2 Spike S1 recombinant protein emulsified with Freund's complete adjuvant. 127 Subsequently, three boosters were given at 14-day intervals emulsified with Freund's 128 incomplete adjuvant. The rabbit was bled and serum was separated on the 5 th day after every 129 booster to determine the Ab titre by Enzyme-Linked Immunosorbent Assay (ELISA). Anti-130 spike S1 antibody was purified using protein-A Sepharose following the product catalogue 149 To further screen the specificity and binding of SARS-CoV-2 Spike S1 antibody, binding 150 ELISA was performed in 96-well microtiter plates. ELISA plates were coated with 0.5 µg/mL 151 SARS-CoV-2 Spike S1 antigen prepared in carbonate buffer (pH 9.6) and incubated overnight 152 at 4 °C. Plate was washed with PBST (0.01%) and blocked for 1 h at 37 °C using 2 % PBSM 153 J o u r n a l P r e -p r o o f SARS-CoV-2 Spike S1 Ab and incubated for 2 h at 37 °C. Plate was washed and 1:10,000 155 dilution of secondary antibody solution was added (HRP labelled anti rabbit anti-IgG Ab) for 156 1 h at 37 °C. Plate was thoroughly washed as described previously and 3,3′,5,5′- 179 Various physicochemical methods were used to confirm the labelling of SARS-CoV-2 Ab with Competitive ELISA titration curve to determine LOD of SARS-CoV-2 Spike S1 Ag with in-242 house developed SARS-CoV-2 Spike S1 Ab. 245 Figure 2 A showed the characteristic peak of AuNPs at 520 nm due to its surface plasmon 246 resonance (SPR) properties whereas a red shift of 9 nm was observed at 529 nm when AuNPs 247 were labelled with SARS-CoV-2 Ab due to an increase in the size of the complex that 305 Both DPV and CV were used for the determination of SARS-CoV-2 spike S1 Ag concentration 306 as shown in Figure 4 A and C. With increasing SARS-CoV-2 spike S1 Ag concentration, the In this proposed work, we have successfully developed FTO/AuNPs/SARS-CoV-2Ab Development of a 529 common biosensor format for an enzyme based biosensor array to monitor fruit 530 quality Detection of Cancer-Specific Proteases Using 532 Magnetic Relaxation of Peptide-Conjugated Nanoparticles in Biological Environment Microfluidic paper device for rapid detection of aflatoxin B1 using an aptamer based 536 colorimetric assay Graphene nanosheets as an electric mediator for 538 ultrafast sensing of urokinase plasminogen activator receptor-A biomarker of cancer A smart nanosensor for the detection of human immunodeficiency virus and 542 associated cardiovascular and arthritis diseases using functionalized graphene-based 543 transistors Electrochemical Sensors Gold nanoparticle-based 546 electrochemical biosensors Graphene based sensors Investigation of Electrical Conductivity of Gold Nanoparticles Scattered in 551 Polyvinylidene Fluoride/Polyvinyl Chloride via Laser Ablation for Electrical 552 Influence of Nanoparticles on 554 Thermal and Electrical Conductivity of Composites Spectrometric Identification of SARS-CoV-2 Proteins from Gargle Solution Samples 588 of COVID-19 Patients An ultrasensitive, rapid, and portable 590 coronavirus SARS-CoV-2 sequence detection method based on CRISPR-Cas12 Cas12-based detection of SARS-CoV-2 Ultra-sensitive and high-throughput CRISPR-p owered COVID-19 598 diagnosis A Single and Two-Stage, Closed-Tube, Molecular 600 Test for the 2019 Novel Coronavirus (COVID-19) at Home, Clinic, and Points of 601 Entry RT-LAMP for rapid diagnosis of 604 coronavirus SARS-CoV-2 Development of a reverse transcription-loop-mediated isothermal 608 amplification as a rapid early-detection method for novel SARS-CoV-2 Rapid and 612 extraction-free detection of SARS-CoV-2 from saliva with colorimetric LAMP Dual-Functional Serum Test for SARS-CoV-2 Antibodies with Portable Surface Plasmon Resonance 620 Sensing Selective Naked-Eye Detection of SARS-CoV-2 Mediated by N Gene 623 Targeted Antisense Oligonucleotide Capped Plasmonic Nanoparticles