Abstract
In this study, we have developed a platform for the rapid screening of the SARS-CoV-2 virus in saliva. Electrical double layer (EDL) gated field-effect transistors (FET) biosensor is applied in this research to detect the electrical signals in the sample. Nucleocapsid protein is used as the target protein since it is expressed abundantly during infection. The sensor illustrates a lower detection limit compared to recent methods, which shows the potential of our sensor to make a diagnosis in the early stages of the disease. Besides, additional pre-pretreatment is not needed. The sensor is a robust diagnostic tool for rapid screening of COVID-19. Introduction The coronavirus pandemic, also known as COVID-19, is caused by SARS-CoV-2 virus. COVID-19 causes severe acute respiratory syndrome and poses a significant threat to health internationally. The World Health Organization (WHO) declared this as a global pandemic. Symptom-based screening cannot be an effective strategy to identify individuals. The symptoms of mild patients are similar to influenza, which may easily lead to false negative confirmation of disease. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR) is the most well-known method employed for the clinical diagnosis of COVID-19. But, this method is costly and needs a long time to get the result. Hence, a rapid and low-cost diagnosis method is urgently required. Experimental Sensor array fabrication The disposable sensor array chip consists of two gold electrodes. One electrode is connected to the gate voltage supply, defined as the reference electrode. The other one is connected to the gate metal, and considered as the sensing electrode. Functionalization of sensor surface Antibody against SARS-CoV-2 nucleocapsid protein is used as receptor in this study, which is immobilized on the gold gate electrode. Traut’s Reagent (Thermo Scientific, (26101)) introduces sulfhydryl (-SH) groups to the primary amines of antibody. The modified antibody can then covalently bind to Au gate electrode by Au-S Bond. The antibody(1.5mg/mL) and Traut’s reagent (14 mM) are mixed and incubated at room temperature for 1 hour. The mixture is then dropped on the sensor surface and incubated in 4 °C refrigerator for 12 hours. Electrical measurement The sensor array chip is inserted into a portable prototype device with FET LND150. 2V DC bias as the drain-source voltage and a short duration gate pulse are steadily provided. The difference between the two current drains is defined as current gain, which is used as the index in data analysis. Fluorescence experiment After the electrical measurement, the secondary antibody, anti-mouse IgG (DyLight® 594), is added to the sensor to conjugate with the primary antibody. The sensor array chip is incubated for 1 hour at room temperature, and the unbound molecule is washed away by 1X PBS. Fluorescence microscope, (LEICA DM2500 LED) is used to capture the fluorescence image with 100 ms exposure time and quantitative analysis is performed using imageJ software. Result and conclusion When infected with SARS-CoV-2, the body produces antibodies that bind specific to nucleocapsid proteins and other surface antigens to help eliminate the virus. SARS-CoV-2 virus nucleocapsid protein is abundantly expressed during infection and is one of the highly immunogenic proteins. Hence, nucleocapsid protein is used as the target protein in this study. We immobilized antibody for nucleocapsid protein on the sensor. Electrical signal changed as the concentration of nucleocapsid protein increased from 0 to 400 ng/mL. The limit of detection for N protein was established at 0.4ng/mL. Tests that can distinguish between IgM and IgG can provide information about the stage of infection, indicating how long the person has been infected with SARS-CoV-2. In this study, we developed a platform that can help to confirm the infection in patients quickly in saliva. N-protein testing can potentially help us track the spread of the disease and accurately detect the infected individuals so as to control further spreading of the disease. Figure 1
Published Version
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