ABSTRACT
More than 6,000,000 people have died due to the coronavirus (COVID-19) pandemic. This disease spread quickly due to its highly contagious nature. The SARS-CoV-2 virus that causes the disease can be transmitted through saliva droplets secreted by infected people at a distance of less than 1 m. As a result, saliva has been accepted as an alternative specimen for COVID-19 detection by the Centers for Disease Control and Prevention (CDC). Furthermore, WHO recommended the use of rapid antigen tests based on lateral flow immunoassay when reverse transcription-polymerase chain reaction (RT-PCR) is not available. We developed a saliva-based rapid antigen test by optimizing the antibody concentration and optimum pH for the conjugation of antibody and gold nanoparticles. We found that the best running buffer formulation consisted of 75 mM sodium phosphate buffer, 1% NaCl, 1% Triton X-100, 0.5% N-acetyl-L-cysteine, and 0.02% sodium azide. The addition of a mucolytic agent in the buffer can reduce the viscosity of saliva, thus improving sensitivity. The rapid test developed detected the lowest concentration of nucleocapsid protein at 0.1 μg/mL. Our study revealed 100% specificity against negative COVID-19 saliva and no cross-reaction with avian influenza virus hemagglutinin. © 2022 THE AUTHOR(S). This article is distributed under a Creative Commons Attribution-ShareAlike 4.0 International license.
ABSTRACT
SAR S disease reappeared at the end of 2019 with a new name as Coronavirus Disease 2019 (COVID-19) caused by a new virus called SARS-CoV-2. This virus has spread throughout the world until recently and caused massive deaths and losses. The nucleic acid test in the form of real-time reverse transcriptase-polymerase chain reaction (RT-PCR) is very important to diagnose COVID-19 in patients, but this method has several drawbacks such as operators who have to be trained, the diagnosis results appear in a relatively long time, and the examination price relatively expensive. This research was conducted to produce immunoglobulin Y (IgY) extracted from chicken egg yolk targeting the S-protein receptor-binding domain (RBD) on SARSCoV- 2 as a component of the surface plasmon resonance (SPR) SARS-CoV-2 antigen detection kit. This research was started by extracting IgY from hyperimmune chicken egg yolks with the polyethylene glycol (PEG) precipitation method and continued with dialysis. The extracted IgY was further purified using thiophilic adsorption chromatography and concentrated by using Amicon® Ultra-15 ultrafiltration. The IgY activity against SARS-CoV-2 RBD was tested qualitatively using the agar gel precipitation test (AGPT) technique and the total protein content was determined using the Lowry method. IgY was tested for its affinity against SARS-CoV-2 RBD using SPR. The IgY concentration obtained was 11 mg/mL. The AGPT result showed the presence of IgY activity against SAR S-CoV-2 RBD isolated from egg yolk and chicken serum after 8 weeks after the first vaccination of chickens. The SDS-PAGE results showed a very clear band of IgY characters. The obtained IgY showed adequate interaction with commercial SARS-CoV-2 RBD on an SPR device. The purified IgY was able to bind with protein-S RBD and showed a fairly good affinity for the SARS-CoV-2 antigen sample. The results of these observations indicate that IgY anti-S-protein SARS-CoV-2 can be produced and purified from chicken egg yolk and can be used as a diagnostic component to detect SARS-CoV-2 antigen, especially on SPR.