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Single-molecule fluorescence resonance energy transfer (smFRET) requires significant spectral overlap between the donor emission and the acceptor absorption, and its efficiency is strongly dependent on the donor-acceptor distance. Consequently, smFRET has the capability to study molecular interactions of biomolecules with sub-nanometer resolution in a homogeneous manner without the involvement of any separation steps. The smFRET has significant advantages of high signal-to-noise ratio, low sample consumption, and homogeneous assay, providing a new approach to detect low-abundant biomolecules with extremely high sensitivity. In this review, we summarize the strategies for improving smFRET efficiency and the applications of smFRET in sensitive detection of DNAs, microRNAs, enzymes, and in situ imaging of single-nucleotide mutation. Moreover, we provide new insight into the challenges and future perspectives of smFRET. • Single-molecule fluorescence resonance energy transfer (smFRET) provides a new approach to detect low-abundant biomolecules. • We summarize the strategies for improving smFRET efficiency. • We review the applications of smFRET in sensitive detection of DNAs, microRNAs and enzymes. • We provide new insight into the challenges and future perspectives of smFRET. [ABSTRACT FROM AUTHOR]

• Innovates miRNA detection with cholesterol-enhanced 3D DNA Walker, enhancing precision. • Uses micelle structure for nuclease protection, cutting false positives in samples. • Allows cancer cell autonomous miRNA imaging, no need for external cofactors. • Enables precise cancer subtype identification via multiple miRNAs, boosting accuracy. • Advances molecular diagnostics, key for early detection and personalized medicine. Achieving reliable miRNA detection and imaging remains a critical and formidable challenge. To address this challenge, we developed a three-dimensional (3D) DNA walker nanoprobe (DWN), derived from a multifunctional molecular beacon with a lipidic moiety. This innovative structure employs a cholesterol-conjugated multifunctional molecular beacon (Chol-MMB) assembled through hydrophobic interactions, thereby bypassing the cumbersome and inefficient process of immobilizing DNA strands onto rigid surfaces. Once the walker nanoprobe enters cells, the binding between the target miRNA and the nanoprobe triggers an intermolecular polymerization process, leading to a conformational change in DWN. This change liberates a walking strand, initiating the walking reaction. As a result, the 3D DNA walker nanoprobe illuminates the fluorophore, enabling precise quantitative analysis. The efficacy of the DWN was demonstrated by the limit of detection of 0.326 fmol for miR-21 with a reaction duration of 1 h. In addition, this strategy effectively differentiates various cells based on the specific miRNA. Further refinement of this method will lead to an efficacious miRNA-based platform for cell typing and liquid biopsy. [ABSTRACT FROM AUTHOR]

Introduction Myocardial ischemia-reperfusion (I/R) injury affects millions of people worldwide and has a very high mortality rate. Since microRNA-497 (miR-497) has been found to be related with cardiomyocyte apoptosis, this study aimed to explore the effect of miR-497 by targeting Mfn2 in a mouse model of myocardial ischemia-reperfusion (I/R) injury. Materials BALB/c mice were modeled with I/R and some were injected with miR-497 agomir before I/R to observe whether miR-497 alleviates the injury that occurs as a result of I/R. Bioinformatics website and dual-luciferase reporter gene assay were employed in order to detect the relations between miR-497 and Mfn2 gene. Next, cells were extracted to be transfected with different mimic, inhibitor and siRNAs to further explore how miR-497 acts to I/R. Western blot analysis and reverse transcription quantitative polymerase chain reaction (RT-qPCR) were conducted to measure expressions of miR-497, Mfn2, Fas, Bcl-2, Bax and Caspase-3 in myocardial tissues and cardiomyocytes after transfection. CCK-8 assay and flow cytometry were used to determine proliferation, cell cycle distribution and apoptosis of cardiomyocytes in each group after transfection. Results Mice with I/R had myocardial dysfunction but before the injection with miR-497 agomir, the impairment was alleviated. Mfn2 was verified as the target gene of miR-497. The inhibition of miR-497 in turn inhibits Mfn2 expressione and cardiomyocyte apoptosis. The overexpression of miR-497 and Mfn2 gene silencing can lead to the promotion of proliferation capability of mice cardiomyocytes in vitro. Overexpressed miR-497 and Mfn2 gene silencing can also facilitate cell cycle entry and inhibit the apoptosis cardiomyocytes of mice in vitro. Conclusion The present study provided strong evidence that miR-497 promotes proliferation and inhibits apoptosis of cardiomyocytes by downregulating the expression of Mfn2 in a mouse model of myocardial I/R injury. [ABSTRACT FROM AUTHOR]