Fluorescence intensity coded DNA frameworks based on the FRET effect enable multiplexed miRNA imaging in living cells

Area of Science:

• Biochemistry
• Molecular Biology
• Nanotechnology

Background:

• MicroRNA (miRNA) analysis is crucial for cancer diagnosis, treatment, and prognosis.
• Multiplexed miRNA imaging offers enhanced diagnostic and prognostic capabilities.
• Existing imaging strategies require novel approaches for improved accuracy and multiplexing.

Purpose of the Study:

• To develop a novel fluorescence emission intensity (FEI) encoding strategy for multiplexed miRNA imaging.
• To utilize a tetrahedron DNA framework (TDF) carrier with Förster resonance energy transfer (FRET) between Cy3 and Cy5.
• To demonstrate the application of this strategy for intracellular multiplexed miRNA detection.

Main Methods:

• Construction of six FEI-encoded TDF (FEI-TDF) samples by varying Cy3 and Cy5 labeling on the TDF.
• Characterization of distinct FEIs and spectral properties of FEI-TDF samples in vitro.
• Improvement of FEI stability by dividing spectral ranges.
• Development of five distinct codes based on FEI ranges.
• Assessment of TDF biocompatibility using CCK-8 assay.
• Design of barcode probes for multiplexed miRNA imaging in MCF-7 cells.

Main Results:

• Distinct FEIs and colors were observed for FEI-TDF samples under UV irradiation.
• Improved stability of FEIs was achieved by spectral range division.
• Five highly discriminative codes were developed based on FEI ranges.
• TDF carrier demonstrated excellent biocompatibility.
• Successful multiplexed imaging of miRNA-16, miRNA-21, and miRNA-10b in MCF-7 cells was achieved with distinct merged colors.

Conclusions:

• The developed FEI-TDF strategy offers a novel approach for fluorescence multiplexing.
• This method provides a new perspective for developing advanced fluorescence multiplexing strategies.
• The strategy shows potential for precise diagnosis and prognosis in cancer through enhanced miRNA imaging.