Gene

SIGNAL (programming language)

microRNA

Cell biology

Biology

Molecular probe

Materials science

DNA replication

DNA extraction

Literature

Biochemistry

Molecule

Computer science

In situ

Fluorescence-lifetime imaging microscopy

Rolling circle replication

Physics

Nanotechnology

Organic chemistry

Optics

Fluorescence

Multiple displacement amplification

Chemistry

Programming language

Dual (grammatical number)

Colocalization

Polymerase chain reaction

Biophysics

Biomolecule

Optoelectronics

ACS Nano

The in vitro detection of low abundance biomolecules via nonenzymatic signal amplification is an attractive strategy. However, it remains a challenge to monitor targets of interest in situ in living cells by low-background interference and visualized enzyme-free signal amplification strategies. Taking advantage of the single-molecule imaging and dynamic DNA nanotechnologies, we have achieved the target-triggered self-assembly of nanostructure-based dual-color fluorescent probes (NDFPs) by an enzyme-free toehold-mediated strand displacement cascade. NDFPs will facilitate the simple and visualized monitoring of microRNA (miRNA) at the femtomolar level. The recycled miRNA can be considered as the catalyst for the assembly of multiple H1/H2 duplexes. This generated the fluorescence signal of the enhanced target expression, indicating both in vitro and in vivo signal-amplified imaging. Moreover, the NDFPs improved the measurement accuracy by dual-color colocalization imaging to greatly avoid false-positive signals and enabled the successful in situ imaging of miRNA in living cells in real time. This work provides a strategy to visually monitor and study the integration of signal amplification detection and single-molecule imaging. NDFPs may be an important step toward the enzyme-free amplified monitoring and imaging of various biomolecules in living cells at the single-molecule level.

Construction of Dual-Color Probes with Target-Triggered Signal Amplification for <i>In Situ</i> Single-Molecule Imaging of MicroRNA

Construction of Dual-Color Probes with Target-Triggered Signal Amplification for In Situ Single-Molecule Imaging of MicroRNA