Transient Hybridization Directed Nanoflare for Single-Molecule miRNA Imaging.
Lina LiYingjie YuCongshan WangQianqian HanXin SuPublished in: Analytical chemistry (2019)
Accurate quantifications of cellular miRNAs are important not only for accelerating them becoming reliable diagnostics biomarkers but also for deeply understanding their influence on central signaling pathways. Although single-molecule miRNA imaging permits quantifying biomolecules at the single-molecule level, it is limited by the sensitivity and specificity of hybridization-based probes. We report a miRNA single-molecule imaging method by using conjugated polymer nanoparticle (CPN) labeled short DNA probe termed as a nanoflare. The transient hybridization of the nanoflares and target miRNAs yields a featured single-molecule kinetics signal rendering high single-molecule sensitivity and specificity. miRNA can be detected with a remarkable detection limit of 1 fM without using any amplification steps. The discrimination capability of homologous miRNAs was also demonstrated. Taking advantage of the featured single-molecule signal of the nanoflare, we can directly count single miR-21 molecules in single cells by using highly inclined and laminated optical sheet (HILO) microscopy. The statistics of the counting reveals miR-21's cell-to-cell fluctuation and differential expression of tumor cells and normal cells.
Keyphrases
- single molecule
- living cells
- high resolution
- atomic force microscopy
- induced apoptosis
- cell proliferation
- long non coding rna
- signaling pathway
- single cell
- cell therapy
- mass spectrometry
- dna damage
- long noncoding rna
- dna repair
- endoplasmic reticulum stress
- stem cells
- epithelial mesenchymal transition
- quantum dots
- bone marrow
- nucleic acid
- peripheral blood
- mesenchymal stem cells
- pet imaging
- sensitive detection
- structural basis