Evaluation of a lyophilized CRISPR-Cas12 assay for a sensitive, specific, and rapid detection of SARS-CoV-2
We evaluated a lyophilized CRISPR-Cas12 assay for SARS-CoV-2 detection (Lyo-CRISPR SARS-CoV-2 kit) based on reverse transcription, isothermal amplification, and CRISPR-Cas12 reaction. From a total of 210 RNA samples extracted from nasopharyngeal swabs using spin columns, the Lyo-CRISPR SARS-CoV-2 ki...
| Autores principales: | , , , , , , , , , , , , , , |
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| Formato: | Artículo |
| Lenguaje: | Inglés |
| Publicado: |
MDPI
2021
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| Materias: | |
| Acceso en línea: | http://hdl.handle.net/20.500.12123/9128 https://www.mdpi.com/1999-4915/13/3/420 https://doi.org/10.3390/v13030420 |
| _version_ | 1855484336657137664 |
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| author | Curti, Lucía Ana Primost, Ivana Valla, Sofia Ibañez Alegre, Daiana Olguin Perglione, Cecilia Repizo, Guillermo Daniel Lara, Julia Parcerisa, Ivana Palacios, Antonela Llases, María Eugenia Rinflerch, Adriana Barrios, Melanie Pereyra Bonnet, Federico Gimenez, Carla Alejandra Marcone, Débora Natalia |
| author_browse | Barrios, Melanie Curti, Lucía Ana Gimenez, Carla Alejandra Ibañez Alegre, Daiana Lara, Julia Llases, María Eugenia Marcone, Débora Natalia Olguin Perglione, Cecilia Palacios, Antonela Parcerisa, Ivana Pereyra Bonnet, Federico Primost, Ivana Repizo, Guillermo Daniel Rinflerch, Adriana Valla, Sofia |
| author_facet | Curti, Lucía Ana Primost, Ivana Valla, Sofia Ibañez Alegre, Daiana Olguin Perglione, Cecilia Repizo, Guillermo Daniel Lara, Julia Parcerisa, Ivana Palacios, Antonela Llases, María Eugenia Rinflerch, Adriana Barrios, Melanie Pereyra Bonnet, Federico Gimenez, Carla Alejandra Marcone, Débora Natalia |
| author_sort | Curti, Lucía Ana |
| collection | INTA Digital |
| description | We evaluated a lyophilized CRISPR-Cas12 assay for SARS-CoV-2 detection (Lyo-CRISPR SARS-CoV-2 kit) based on reverse transcription, isothermal amplification, and CRISPR-Cas12 reaction. From a total of 210 RNA samples extracted from nasopharyngeal swabs using spin columns, the Lyo-CRISPR SARS-CoV-2 kit detected 105/105 (100%; 95% confidence interval (CI): 96.55–100) positive samples and 104/105 (99.05%; 95% CI: 94.81–99.97) negative samples that were previously tested using commercial RT-qPCR. The estimated overall Kappa index was 0.991, reflecting an almost perfect concordance level between the two diagnostic tests. An initial validation test was also performed on 30 nasopharyngeal samples collected in lysis buffer, in which the Lyo-CRISPR SARS-CoV-2 kit detected 20/21 (95.24%; 95% CI: 76.18–99.88) positive samples and 9/9 (100%; 95% CI: 66.37–100) negative samples. The estimated Kappa index was 0.923, indicating a strong concordance between the test procedures. The Lyo-CRISPR SARS-CoV-2 kit was suitable for detecting a wide range of RT-qPCR-positive samples (cycle threshold range: 11.45–36.90) and dilutions of heat-inactivated virus (range: 2.5–100 copies/µL); no cross-reaction was observed with the other respiratory pathogens tested. We demonstrated that the performance of the Lyo-CRISPR SARS-CoV-2 kit was similar to that of commercial RT-qPCR, as the former was highly sensitive and specific, timesaving (1.5 h), inexpensive, and did not require sophisticated equipment. The use of this kit would reduce the time taken for diagnosis and facilitate molecular diagnosis in low-resource laboratories. |
| format | Artículo |
| id | INTA9128 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2021 |
| publishDateRange | 2021 |
| publishDateSort | 2021 |
| publisher | MDPI |
| publisherStr | MDPI |
| record_format | dspace |
| spelling | INTA91282021-04-19T17:48:58Z Evaluation of a lyophilized CRISPR-Cas12 assay for a sensitive, specific, and rapid detection of SARS-CoV-2 Curti, Lucía Ana Primost, Ivana Valla, Sofia Ibañez Alegre, Daiana Olguin Perglione, Cecilia Repizo, Guillermo Daniel Lara, Julia Parcerisa, Ivana Palacios, Antonela Llases, María Eugenia Rinflerch, Adriana Barrios, Melanie Pereyra Bonnet, Federico Gimenez, Carla Alejandra Marcone, Débora Natalia CRISPR Severe Acute Respiratory Syndrome Coronavirus 2 COVID-19 Diagnosis Fluorescence Freeze Drying Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Interespaciadas Coronavirus del Síndrome Respiratorio Agudo Grave 2 Diagnóstico Fluorescencia Liofilización SARS-CoV-2 We evaluated a lyophilized CRISPR-Cas12 assay for SARS-CoV-2 detection (Lyo-CRISPR SARS-CoV-2 kit) based on reverse transcription, isothermal amplification, and CRISPR-Cas12 reaction. From a total of 210 RNA samples extracted from nasopharyngeal swabs using spin columns, the Lyo-CRISPR SARS-CoV-2 kit detected 105/105 (100%; 95% confidence interval (CI): 96.55–100) positive samples and 104/105 (99.05%; 95% CI: 94.81–99.97) negative samples that were previously tested using commercial RT-qPCR. The estimated overall Kappa index was 0.991, reflecting an almost perfect concordance level between the two diagnostic tests. An initial validation test was also performed on 30 nasopharyngeal samples collected in lysis buffer, in which the Lyo-CRISPR SARS-CoV-2 kit detected 20/21 (95.24%; 95% CI: 76.18–99.88) positive samples and 9/9 (100%; 95% CI: 66.37–100) negative samples. The estimated Kappa index was 0.923, indicating a strong concordance between the test procedures. The Lyo-CRISPR SARS-CoV-2 kit was suitable for detecting a wide range of RT-qPCR-positive samples (cycle threshold range: 11.45–36.90) and dilutions of heat-inactivated virus (range: 2.5–100 copies/µL); no cross-reaction was observed with the other respiratory pathogens tested. We demonstrated that the performance of the Lyo-CRISPR SARS-CoV-2 kit was similar to that of commercial RT-qPCR, as the former was highly sensitive and specific, timesaving (1.5 h), inexpensive, and did not require sophisticated equipment. The use of this kit would reduce the time taken for diagnosis and facilitate molecular diagnosis in low-resource laboratories. Instituto de Virología Fil: Curti, Lucía Ana. CASPR Biotech; Estados Unidos Fil: Primost, Ivana. Hospital Municipal de Trauma y Emergencias Dr. Federico Abete. Genetics and Molecular Biology Laboratory; Argentina Fil: Valla, Sofia. Universidad Nacional del Noroeste de la Provincia de Buenos Aires. Centro de Investigaciones y Transferencia del Noroeste de la Provincia de Buenos Aires (CITNOBA). Centro de Investigaciones Básicas y Aplicadas (CIBA); Argentina Fil: Valla, Sofia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Ibañez Alegre, Daiana. Universidad Nacional de Misiones. Instituto de Biología Subtropical. Laboratorio Grupo de Investigación en Genética Aplicada (GIGA); Argentina Fil: Ibañez Alegre, Daiana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Olguin Perglione, Cecilia. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Virología; Argentina Fil: Olguin Perglione, Cecilia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Repizo, Guillermo Daniel. CASPR Biotech; Estados Unidos Fil: Lara, Julia. CASPR Biotech; Estados Unidos Fil: Parcerisa, Ivana. CASPR Biotech; Estados Unidos Fil: Palacios, Antonela. CASPR Biotech; Estados Unidos Fil: Llases, María Eugenia. CASPR Biotech; Estados Unidos Fil: Rinflerch, Adriana. Universidad Nacional de Misiones. Instituto de Biología Subtropical. Laboratorio Grupo de Investigación en Genética Aplicada (GIGA); Argentina Fil: Rinflerch, Adriana. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Barrios, Melanie. Universidad de Buenos Aires. Instituto de Producción Agropecuaria; Argentina Fil: Pereyra Bonnet, Federico. CASPR Biotech; Estados Unidos Fil: Gimenez, Carla Alejandra. CASPR Biotech; Estados Unidos Fil: Marcone, Débora Natalia. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Departamento de Microbiología, Inmunología, Biotecnología y Genética. Cátedra de Virología; Argentina Fil: Marcone, Débora Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina 2021-04-19T17:40:04Z 2021-04-19T17:40:04Z 2021-03 info:ar-repo/semantics/artículo info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion http://hdl.handle.net/20.500.12123/9128 https://www.mdpi.com/1999-4915/13/3/420 1999-4915 https://doi.org/10.3390/v13030420 eng info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by-nc-sa/4.0/ Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0) application/pdf MDPI Viruses 13 (3) : 420 (Marzo 2021) |
| spellingShingle | CRISPR Severe Acute Respiratory Syndrome Coronavirus 2 COVID-19 Diagnosis Fluorescence Freeze Drying Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Interespaciadas Coronavirus del Síndrome Respiratorio Agudo Grave 2 Diagnóstico Fluorescencia Liofilización SARS-CoV-2 Curti, Lucía Ana Primost, Ivana Valla, Sofia Ibañez Alegre, Daiana Olguin Perglione, Cecilia Repizo, Guillermo Daniel Lara, Julia Parcerisa, Ivana Palacios, Antonela Llases, María Eugenia Rinflerch, Adriana Barrios, Melanie Pereyra Bonnet, Federico Gimenez, Carla Alejandra Marcone, Débora Natalia Evaluation of a lyophilized CRISPR-Cas12 assay for a sensitive, specific, and rapid detection of SARS-CoV-2 |
| title | Evaluation of a lyophilized CRISPR-Cas12 assay for a sensitive, specific, and rapid detection of SARS-CoV-2 |
| title_full | Evaluation of a lyophilized CRISPR-Cas12 assay for a sensitive, specific, and rapid detection of SARS-CoV-2 |
| title_fullStr | Evaluation of a lyophilized CRISPR-Cas12 assay for a sensitive, specific, and rapid detection of SARS-CoV-2 |
| title_full_unstemmed | Evaluation of a lyophilized CRISPR-Cas12 assay for a sensitive, specific, and rapid detection of SARS-CoV-2 |
| title_short | Evaluation of a lyophilized CRISPR-Cas12 assay for a sensitive, specific, and rapid detection of SARS-CoV-2 |
| title_sort | evaluation of a lyophilized crispr cas12 assay for a sensitive specific and rapid detection of sars cov 2 |
| topic | CRISPR Severe Acute Respiratory Syndrome Coronavirus 2 COVID-19 Diagnosis Fluorescence Freeze Drying Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Interespaciadas Coronavirus del Síndrome Respiratorio Agudo Grave 2 Diagnóstico Fluorescencia Liofilización SARS-CoV-2 |
| url | http://hdl.handle.net/20.500.12123/9128 https://www.mdpi.com/1999-4915/13/3/420 https://doi.org/10.3390/v13030420 |
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