Silica bead-based microfluidic device with integrated photodiodes for the rapid capture and detection of rolling circle amplification products in the femtomolar range.
Biosens Bioelectron
; 128: 68-75, 2019 Mar 01.
Article
en En
| MEDLINE
| ID: mdl-30634076
ABSTRACT
The rapid and sensitive detection of specific nucleic acid sequences at the point-of-care (PoC) is becoming increasingly in demand for a variety of emergent biomedical applications ranging from infectious disease diagnostics to the screening of antimicrobial resistance. To meet such demand, considerable efforts have been invested towards the development of portable and integrated analytical devices combining microfluidics with miniaturized signal transducers. Here, we demonstrate the combination of rolling circle amplification (RCA)-based nucleic acid amplification with an on-chip size-selective trapping of amplicons on silica beads (~8 nL capture chamber) coupled with a thin-film photodiode (200â¯×â¯200⯵m area) fluorescence readout. Parameters such as the flow rate of the amplicon solution and trapping time were optimized as well as the photodiode measurement settings, providing minimum detection limits below 0.5 fM of targeted nucleic acids and requiring only 5⯵L of pre-amplified sample. Finally, we evaluated the analytical performance of our approach by benchmarking it against a commercial instrument for RCA product (RCP) quantification and further investigated the effect of the number of RCA cycles and elongation times (ranging from 10 to 120â¯min). Moreover, we provide a demonstration of the application for diagnostic purposes by detecting RNA from influenza and Ebola viruses, thus highlighting its suitability for integrated PoC systems.
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Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
ARN
/
Técnicas Biosensibles
/
Ebolavirus
Tipo de estudio:
Diagnostic_studies
Idioma:
En
Revista:
Biosens Bioelectron
Asunto de la revista:
BIOTECNOLOGIA
Año:
2019
Tipo del documento:
Article
País de afiliación:
Portugal