"Paper Machine" for Molecular Diagnostics.
Anal Chem
; 87(15): 7595-601, 2015 Aug 04.
Article
em En
| MEDLINE
| ID: mdl-26104869
ABSTRACT
Clinical tests based on primer-initiated amplification of specific nucleic acid sequences achieve high levels of sensitivity and specificity. Despite these desirable characteristics, these tests have not reached their full potential because their complexity and expense limit their usefulness to centralized laboratories. This paper describes a device that integrates sample preparation and loop-mediated isothermal amplification (LAMP) with end point detection using a hand-held UV source and camera phone. The prototype device integrates paper microfluidics (to enable fluid handling) and a multilayer structure, or a "paper machine", that allows a central patterned paper strip to slide in and out of fluidic path and thus allows introduction of sample, wash buffers, amplification master mix, and detection reagents with minimal pipetting, in a hand-held, disposable device intended for point-of-care use in resource-limited environments. This device creates a dynamic seal that prevents evaporation during incubation at 65 °C for 1 h. This interval is sufficient to allow a LAMP reaction for the Escherichia coli malB gene to proceed with an analytical sensitivity of 1 double-stranded DNA target copy. Starting with human plasma spiked with whole, live E. coli cells, this paper demonstrates full integration of sample preparation with LAMP amplification and end point detection with a limit of detection of 5 cells. Further, it shows that the method used to prepare sample enables concentration of DNA from sample volumes commonly available from fingerstick blood draw.
Texto completo:
1
Temas:
ECOS
/
Aspectos_gerais
Bases de dados:
MEDLINE
Assunto principal:
Papel
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Técnicas de Diagnóstico Molecular
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Técnicas Analíticas Microfluídicas
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
En
Revista:
Anal Chem
Ano de publicação:
2015
Tipo de documento:
Article
País de afiliação:
Estados Unidos