RESUMEN
We develop a novel amplified split aptamer sensor for highly sensitive detection and imaging of small molecules in living cells by using cationic block copolymer nanoparticles (BCNs) with entrapped fluorescent conjugated polymer as a delivery agent. The design of a split aptamer as the initiator of hybridization chain reaction (HCR) affords the possibility of enhancing the signal-to-background ratio and thus allows high-contrast imaging for small molecules with relatively weak interactions with their aptamers. The novel design of using fluorescent cationic BCNs as the nanocarrier enables efficient and self-tracking transfection of DNA probes. Results reveal that BCNs exhibit high fluorescence brightness allowing direct tracking of the delivery location. The developed amplified split aptamer sensor is shown to have high sensitivity and selectivity for in vitro quantitative detection of adenosine triphosphate (ATP) with a detection limit of 30 nM. Live cell studies show that the sensor provides a "signal on" approach for specific, high-contrast imaging of ATP. The DNA sensor based HCR system may provide a new generally applicable platform for detection and imaging of low-abundance biomarkers.
Asunto(s)
Aptámeros de Nucleótidos/química , Técnicas Biosensibles/métodos , Nanopartículas/química , Polietilenglicoles/química , Polivinilos/química , Adenosina Trifosfato/análisis , Aptámeros de Nucleótidos/genética , Biomarcadores/análisis , ADN/química , ADN/genética , Sondas de ADN/química , Sondas de ADN/genética , Fluorenos/química , Fluorenos/toxicidad , Colorantes Fluorescentes/química , Colorantes Fluorescentes/toxicidad , Células HeLa , Humanos , Secuencias Invertidas Repetidas , Límite de Detección , Microscopía Fluorescente/métodos , Nanopartículas/toxicidad , Hibridación de Ácido Nucleico , Polietilenglicoles/toxicidad , Polivinilos/toxicidadRESUMEN
A novel isothermal nucleic acid amplification technology is developed by coupling catalytic hairpin assembly with enzymatic repairing amplification as a highly sensitive and selective platform for miRNA detection.