Molecular Diagnostics of Dengue by Reverse Transcription-Loop Mediated Isothermal Amplification (RT-LAMP) in Disposable Polyester-Toner Microdevices
| Autor: | Geovana de Melo Mendes, Fabíola Souza Fiaccadori, Célia Maria de Almeida Soares, Alexandre Melo Bailão, Kézia Gomes de Oliveira, Juliane C. Borba, Thaís Santana de Oliveira, Emanuel Carrilho, Gabriela R. M. Duarte, Maurício Lacerda Nogueira |
|---|---|
| Rok vydání: | 2019 |
| Předmět: |
Gel electrophoresis
DENGUE 010401 analytical chemistry Loop-mediated isothermal amplification RNA General Chemistry Dengue virus medicine.disease medicine.disease_cause Molecular diagnostics 01 natural sciences Virology 0104 chemical sciences Dengue fever chemistry.chemical_compound chemistry medicine Reverse Transcription Loop-mediated Isothermal Amplification DNA |
| Zdroj: | Repositório Institucional da USP (Biblioteca Digital da Produção Intelectual) Universidade de São Paulo (USP) instacron:USP |
| Popis: | Dengue is one of the most prevalent infectious tropical diseases in the world, with high incidence in over 100 countries. Rapid and reliable diagnosis of dengue is of great importance for public health. To simplify molecular diagnostics, isothermal amplification techniques have recently emerged as an alternative to conventional methods of deoxyribonucleic acid (DNA) amplification. Here, we developed a one-step method for dengue virus detection from real sample based on RT-LAMP (reverse transcription-loop mediated isothermal amplification) in a disposable microdevice. The reaction was thermally controlled with a thermoblock for 15 min at 72 oC. At the end of the incubation time, we either removed the solution for detection of fragments by gel electrophoresis or added DNA intercalator for visual detection on-chip. Our results demonstrated that it is possible to detect dengue virus through RT-LAMP directly from a serum sample, without previous ribonucleic acid (RNA) extraction. The success of RT-LAMP was confirmed in reactions initiated with 0.8 fg µL-1 of RNA, which represents 200 copies of RNA per µL. RT-LAMP in a polyester-toner (PeT) microdevice is a simple and inexpensive method that allows for rapid detection of dengue virus with high reliability and great potential for point-of-care applications. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|