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Authors
Advisor(s)
Abstract(s)
A pandemia da doença de coronavírus 2019 (COVID-19) veio representar grandes ameaças ao mundo em diversos aspetos. Nesta situação é essencial haver uma testagem regular de toda a população. Já existem vários testes disponíveis para deteção de Síndrome respiratório aguda grave - coronavírus 2 (SARS-CoV-2), no entanto demos especial atenção às técnicas de antigénio e de diagnóstico molecular. Na primeira técnica a amostra foi colocada numa cassete de cromatografia, que possuía uma área revestida com o anticorpo monoclonal anti-SARS-CoV-2. A segunda técnica dividiu-se em dois testes. O primeiro foi o teste VitaPCR™, onde a amostra foi armazenada num tampão específico, que lisou e extraiu a amostra numa única etapa, sendo possível a deteção do SARS-CoV-2 através da fluorescência em múltiplos canais. O segundo teste foi o Reverse Transcription–Polymerase Chain Reaction em tempo real (rRT-PCR). Neste, houve primeiramente a extração do ácido nucleico através do uso de beads magnéticas, seguindo-se para a fase de amplificação através do rRT-PCR que serviu para detetar a presença dos genes S, RdRP e N. Este estágio serviu para realizar todo o processo analítico de amostras com suspeita de SARS-CoV-2. Foi feita uma análise de cada um dos testes de modo a entender os seus princípios teóricos, metodologia, resultados possíveis e respetiva interpretação. Em suma, foi concluído que o teste rRT-PCR é o método mais adequado para o diagnóstico de COVID-19. No entanto, os testes VitaPCR™ e de deteção de antigénio, foram considerados melhores como métodos de Point of Care (POC), prestando um grande auxílio no controlo da pandemia e diminuição da sobrecarga nos hospitais.
ABSTRACT - The coronavirus disease 2019 (COVID-19) pandemic has come to pose major threats to the world in many ways. In this situation, regular testing of the entire population is essential. There are already several tests available for the detection of Severe Acute Respiratory Syndrome - coronavirus 2 (SARS-CoV-2), however, we paid special attention to antigen and molecular diagnostic techniques. In the first technique, the sample was placed in a chromatography cassette, which had an area coated with the anti-SARS-CoV-2 monoclonal antibody. The second technique was divided into two tests. The first was the VitaPCR™ test, where the sample was stored in a specific buffer, which lysed and extracted the sample in a single step, making it possible to detect SARS-CoV-2 through fluorescence in multiple channels. The second test was the real-time Reverse Transcription–Polymerase Chain Reaction (rRT-PCR). In this one, the nucleic acid was first extracted using magnetic beads, followed by the amplification phase through rRT-PCR, which served to detect the presence of the S, RdRP, and N genes. This stage served to carry out the entire analytical process of samples with suspected SARS-CoV-2. An analysis was made of each of the tests in order to understand their theoretical principles, methodology, possible results, and respective interpretation. In summary, it was concluded that the rRT-PCR test is the most suitable method for the diagnosis of COVID-19. However, VitaPCR™ and antigen detection tests were considered better as Point of Care (POC) methods, providing a great help in controlling the pandemic and reducing the burden on hospitals.
ABSTRACT - The coronavirus disease 2019 (COVID-19) pandemic has come to pose major threats to the world in many ways. In this situation, regular testing of the entire population is essential. There are already several tests available for the detection of Severe Acute Respiratory Syndrome - coronavirus 2 (SARS-CoV-2), however, we paid special attention to antigen and molecular diagnostic techniques. In the first technique, the sample was placed in a chromatography cassette, which had an area coated with the anti-SARS-CoV-2 monoclonal antibody. The second technique was divided into two tests. The first was the VitaPCR™ test, where the sample was stored in a specific buffer, which lysed and extracted the sample in a single step, making it possible to detect SARS-CoV-2 through fluorescence in multiple channels. The second test was the real-time Reverse Transcription–Polymerase Chain Reaction (rRT-PCR). In this one, the nucleic acid was first extracted using magnetic beads, followed by the amplification phase through rRT-PCR, which served to detect the presence of the S, RdRP, and N genes. This stage served to carry out the entire analytical process of samples with suspected SARS-CoV-2. An analysis was made of each of the tests in order to understand their theoretical principles, methodology, possible results, and respective interpretation. In summary, it was concluded that the rRT-PCR test is the most suitable method for the diagnosis of COVID-19. However, VitaPCR™ and antigen detection tests were considered better as Point of Care (POC) methods, providing a great help in controlling the pandemic and reducing the burden on hospitals.
Description
Mestrado em Tecnologias Clínico-Laboratoriais
Keywords
COVID-19 SARS-CoV-2 Antigénio rRT-PCR Antigen
Citation
Rodrigues ID. Estágio em biologia molecular: deteção de SARS-CoV-2 em amostras nasofaríngeas com as técnicas de antigénio, RT-PCR e VitaPCR real time PCR [dissertation]. Lisboa: Escola Superior de Tecnologia da Saúde de Lisboa/Instituto Politécnico de Lisboa; 2022.
Publisher
Instituto Politécnico de Lisboa, Escola Superior de Tecnologia da Saúde de Lisboa