Mismatched fluorescent probes with enhanced strand displacement reaction rate for intracellular long noncoding RNA imaging

We design a mismatched fluorescent probe to directly monitor the long noncoding RNA (lncRNA) in living cells. The introduction of mismatched bases in the fluorescent probe greatly enhances the strand...

Hybridization chain reaction enables a unified approach to multiplexed, quantitative, high-resolution immunohistochemistry and in situ hybridization

ABSTRACT RNA in situ hybridization based on the mechanism of the hybridization chain reaction (HCR) enables multiplexed, quantitative, high-resolution RNA imaging in highly autofluorescent samples, including whole-mount vertebrate embryos, thick brain slices and formalin-fixed paraffin-embedded tissue sections. Here, we extend the benefits of one-step, multiplexed, quantitative, isothermal, enzyme-free HCR signal amplification to immunohistochemistry, enabling accurate and precise protein relative quantitation with subcellular resolution in an anatomical context. Moreover, we provide a unified framework for simultaneous quantitative protein and RNA imaging with one-step HCR signal amplification performed for all target proteins and RNAs simultaneously.

Bright, fluorogenic and photostable avidity probes for RNA imaging

Fluorescent light-up aptamers (FLAPs) emerged as valuable tools to visualize RNA, but are mostly limited by poor brightness, low photostability and high fluorescence background. In this study, we combine bivalent (silicon) rhodamine fluorophores with dimeric FLAPs to yield bright and photostable complexes with low picomolar dissociation constants. Our avidity-based approach resulted in extreme binding strength and high fluorogenicity, enabling single mRNA tracking in living cells.

Hybridization chain reaction enables a unified approach to multiplexed, quantitative, high-resolution immunohistochemistry and in situ hybridization

RNA in situ hybridization (RNA-ISH) based on the mechanism of hybridization chain reaction (HCR) enables multiplexed, quantitative, high-resolution RNA imaging in highly autofluorescent samples including whole-mount vertebrate embryos, thick brain slices, and formalin-fixed paraffin-embedded (FFPE) tissue sections. Here, we extend the benefits of 1-step, multiplexed, quantitative, isothermal, enzyme-free HCR signal amplification to immunohistochemistry (IHC), enabling accurate and precise protein relative quantitation with subcellular resolution in an anatomical context. Moreover, we provide a unified framework for simultaneous quantitative protein and RNA imaging with 1-step HCR signal amplification performed for all target proteins and RNAs simultaneously.

Merging DNA Probes with Nanotechnology for RNA Imaging In Vivo

Background: Imaging of RNA in vivo is of great significance for elucidating their biological functions, revealing mechanisms behind disease, and for further diagnosis and treatment. Over the past decade, a variety of DNA-based molecular imaging techniques have been developed for RNA imaging in living cells. Nevertheless, non-invasive imaging of RNA in animals is still limited. Methods: An overview of the literature involving RNA imaging in vivo based on the integration of DNA probes with nanotechnology has been reviewed. Results: Attributed to DNA’s designability of sequences and specificity of recognition, molecular beacon, strand displacement and hybridization chain reaction would confer quick, efficient and specific response to target RNA. Multifunctional nanomaterials provide powerful support for the intracellular delivery of such DNA probes with spatiotemporal control over their sensing function, thereby achieving RNA imaging in vivo. Conclusion: Merging DNA probes with nanotechnology has gained substantial prospects for RNA imaging in vivo, which not only helps us to better elucidate biological functions of RNA, but also provides valuable information for further disease diagnosis and treatment.