Molecular Diagnostics of Dengue by Reverse Transcription-Loop Mediated Isothermal Amplification (RT-LAMP) in Disposable Polyester-Toner Microdevices

Author(s)

Mendes, Geovana M. ^[1] ; Oliveira, Kezia G. ^[1] ; Borba, Juliane C. ^[2] ; Oliveira, Thais S. ^[3] ; Fiaccadori, Fabiola S. ^[3] ; Nogueira, Mauricio L. ^[4] ; Bailao, Alexandre M. ^[5] ; Soares, Celia Maria A. ^[5] ; Carrilho, Emanuel ^{[2, 6]} ; Duarte, Gabriela R. M. ^[1]

Affiliation

^[1] Univ Fed Goias, Inst Quim, BR-74690900 Goiania, Go - Brazil

^[2] Univ Sao Paulo, Inst Quim Sao Carlos, BR-13566590 Sao Carlos, SP - Brazil

^[3] Univ Fed Goias, Inst Patol Trop & Saude Publ, BR-74605050 Goiania, Go - Brazil

^[4] Fac Med Sao Jose do Rio Preto, Dept Doencas Dermatol Infecciosas & Parasitarias, BR-15090000 Sao Jose Do Rio Preto, SP - Brazil

^[5] Univ Fed Goias, Inst Ciencias Biol, BR-74690900 Goiania, Go - Brazil

^[6] Univ Estadual Campinas, Inst Nacl Ciencia & Tecnol Bioanal, BR-13083970 Campinas, SP - Brazil

• Document type: Journal article
• Source: Journal of the Brazilian Chemical Society; v. 30, n. 9, p. 1841-1849, SEP 2019.
• Web of Science Citations: 1

Abstract

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. (AU)

• FAPESP's process: 14/50867-3 - INCT 2014: National Institute of Science and Technology in Bioanalysis
• Grantee: Marco Aurelio Zezzi Arruda
• Support Opportunities: Research Projects - Thematic Grants
• FAPESP's process: 15/17145-7 - A fully integrated polyester-toner (PT) based microchip for genetic analyses: Development and applications
• Grantee: Emanuel Carrilho
• Support Opportunities: Regular Research Grants