Development of kits for the detection of COVID-19 by the RT-PCR multiplex method in real time and colorimetric by RT-LAMP

The outbreak of the disease caused by the new coronavirus (COVID-19) was declared by the World Health Organization (WHO) a Public Health Emergency of International Importance and later as a pandemic. Worldwide, more than 50,000 deaths have been reported and despite worldwide efforts, there is still no vaccine or specific effective treatments. The strategy recommended by WHO to contain the progress of this pandemic consists of social isolation and the rapid identification of cases of infection. Therefore, diagnostic tests are extremely important and a demand of at least 15 million tests has been estimated by the Ministry of Health to supply the needs of Brazil. The virus nucleic acid amplification test (NAATs) using real-time RT-PCR is the diagnosis recommended by the WHO because it has high sensitivity and specificity, being able to detect the infection at the beginning. However, the reagents needed for this test, currently, must be imported and are scarce internationally due to the magnitude of the pandemic. Therefore, this project has as its initial and immediate objective the viability of a test for RT-PCR in real time multiplex totally produced in Brazil. Cellco currently produces 90% of the inputs needed to compose the standard test, being able to quickly validate and establish the complete kit and supply to centers able to perform the test. However, it is known that RT-qPCR tests are designed and require specific equipment and highly trained personnel, which represents another bottleneck in the ability to carry out mass tests. To overcome this other obstacle, we propose to establish a quick and simple test using the RT-LAMP technique. The application of this methodology for diagnostics has been highly explored because it is possible to adapt it to colorimetric tests with the sensitivity and specificity of other tests by nucleic acid amplification. Additionally, the test can be applied directly to clinical samples, without the need for nucleic acid extraction. Cellco already produces the base enzyme of the LAMP methodology, the DNA Polymerase I of Basillus stearothermophilus (Bst), in their native form and mutants that show greater processivity and resistance to common inhibitors in PCR reactions. We therefore propose the development of an RT-LAMP kit, its adaptation for colorimetric testing and making feasible for rapid coronavirus tests. This methodology is based on specific oligonucleotides, making it possible to maintain their sensitivity and specificity even with mutations and different strains of the virus. With this project, we intend, therefore, to quickly meet the national demand for the RT-qPCR standard test and in parallel to develop a rapid test using the RT-LAMP methodology that allows a large number of tests to be carried out to minimize the effects of the outbreak. COVID-19. (AU) its adaptation for colorimetric testing and feasibility for rapid coronavirus tests. This methodology is based on specific oligonucleotides, making it possible to maintain their sensitivity and specificity even with mutations and different strains of the virus. With this project, we intend, therefore, to quickly meet the national demand for the RT-qPCR standard test and in parallel to develop a rapid test using the RT-LAMP methodology that allows a large number of tests to be carried out to minimize the effects of the outbreak. COVID-19. (AU) its adaptation for colorimetric testing and feasibility for rapid coronavirus tests. This methodology is based on specific oligonucleotides, making it possible to maintain their sensitivity and specificity even with mutations and different strains of the virus. With this project, we intend, therefore, to quickly meet the national demand for the RT-qPCR standard test and in parallel to develop a rapid test using the RT-LAMP methodology that allows a large number of tests to be carried out to minimize the effects of the outbreak. COVID-19. (AU) With this project, we intend, therefore, to quickly meet the national demand for the RT-qPCR standard test and in parallel to develop a rapid test using the RT-LAMP methodology that allows a large number of tests to be carried out to minimize the effects of the outbreak. COVID-19. (AU) With this project, we intend, therefore, to quickly meet the national demand for the RT-qPCR standard test and in parallel to develop a rapid test using the RT-LAMP methodology that allows a large number of tests to be carried out to minimize the effects of the outbreak. COVID-19.