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Development of a rapid antigen test for COVID-19 diagnosis

Marco Torelli

Development of a rapid antigen test for COVID-19 diagnosis.

Rel. Lucia Napione, Francesca Frascella, Alessandro Chiado'. Politecnico di Torino, Corso di laurea magistrale in Ingegneria Biomedica, 2021


COVID-19 is a complex viral disease that spread around the world in the early part of the year 2020, becoming a pandemic in March and overloading the health care systems of most of the world's nations. SARS-CoV-2 is the pathogen responsible for the disease and, like other coronaviruses, is composed of 4 structural proteins: nucleocapsid (N) protein, spike (S) glycoprotein, envelope (E) protein, membrane (M) protein. Even in the presence of effective vaccines, it is very important to have a good contact tracing system, able to monitor and control virus circulation and its new dangerous variants. Moreover, in the presence of a high number of infections, the gold standard qRT-PCR test is not able to properly support the requested demand of tests, due to the need for reagents, specialized personnel and laboratory facilities. On the other hand, point of care rapid antigen tests are characterized by a lower cost, a low turnaround time and do not require specialized personnel or laboratory instrumentation. In particular, promising devices are those based on lateral flow immunoassay (LFIA), which consist of a paper-based strip composed by four components: i) a sample pad, , ii) a conjugate pad, iii) a chromatographic membrane, iv) an adsorbent pad. The purpose of the work presented was to develop a hand-made rapid antigen LFIA device for SARS-CoV-2 detection. This biosensor has some different characteristics compared to commercial rapid antigen tests: S glycoprotein is used as target molecule, moreover it is intended to exploit the natural way that SARS-CoV-2 uses to infect target cells, that is by interacting with angiotensin converting enzyme 2 (ACE-2) through the S1 protein subunit. In this work, recombinant human ACE-2 (rhACE-2) was adopted as the S1 subunit detection reagent instead of using a monoclonal antibody. As detection particles, it was decided to use gold nanoshells (GNSs, 150 nm). In order to increase sensitivity, GNSs were preferred to classic 40 nm gold nanoparticles because of the greater signal that these detection labels are able to provide. The choice of rhACE-2 as detection bioreceptor has the advantage of developing a more robust biosensor, as it is potentially able to detect SARS-CoV-2 regardless of new virus variants circulating in the population. For the test and control lines, monoclonal antibodies specific for the S2 subunit of SARS-CoV-2 S glycoprotein and rhACE-2 were chosen, respectively. Test and control line striping, conjugate pad preparation and LF strips assembly were hand made. A careful attention was paid to the selection of strip component materials as well as to each experimental procedure required for the development of the LFIA device. A simplified sample was used and tested, adding recombinant SARS-CoV-2 S protein ectodomain at known concentration in saline. The detection was successful, showing a limit of detection equal to 25 ng/reaction, despite the issues derived from the device manual assembly. Based on these findings, it is expected to continue the project by preparing the strips in an automated fashion and optimizing the biosensor, moving on to more complex samples. The final goal is to use a salivary swab, instead of the classic nasopharyngeal swab. This would make the test much easier to perform, as the nasopharyngeal swab requires trained personnel for collection. Developing an antigen test that is easier to perform may allow testing a larger number of individuals.

Relators: Lucia Napione, Francesca Frascella, Alessandro Chiado'
Academic year: 2020/21
Publication type: Electronic
Number of Pages: 76
Additional Information: Tesi secretata. Fulltext non presente
Corso di laurea: Corso di laurea magistrale in Ingegneria Biomedica
Classe di laurea: New organization > Master science > LM-21 - BIOMEDICAL ENGINEERING
Aziende collaboratrici: UNSPECIFIED
URI: http://webthesis.biblio.polito.it/id/eprint/19624
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