Versatile Multiplex Endogenous RNA Detection with Simultaneous Signal Normalization Using Mesoporous Silica...

Area of Science:

• Biomedical Engineering
• Molecular Diagnostics
• Cancer Research

Background:

• Accurate detection of endogenous tumor-related RNA is crucial for cancer diagnostics.
• Existing live-cell RNA detection methods suffer from low signal output and cell-to-cell variability due to probe uptake differences.

Purpose of the Study:

• To develop a versatile and highly sensitive mRNA/miRNA nanosensor for live-cell RNA detection.
• To incorporate signal amplification and built-in signal normalization for improved accuracy and reliability.
• To enable multiplexed detection of tumor-related RNAs and serve as a noninvasive imaging tool.

Main Methods:

• Designed a nanosensor using dye-loaded mesoporous silica nanoquenchers (qMSNs) capped with antisense oligos (ASOs).
• Achieved "Turn-ON" fluorescence with signal amplification upon target binding.
• Integrated glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNA-responsive molecular beacons (MBs) for signal normalization.

Main Results:

• Demonstrated successful differentiation between noncancer and cancer cells.
• Accurately monitored relative expression levels of multiple tumor-related RNAs simultaneously in cancer cell lines.
• Achieved high specificity and sensitivity, functioning as a live-cell "qPCR mimic" imaging tool.

Conclusions:

• The developed nanosensor offers a significant advancement in live-cell RNA detection for cancer diagnostics.
• The technology provides a versatile platform for multiplexed RNA detection with built-in normalization.
• This nanosensor enables noninvasive, quantitative monitoring of tumor RNA expression in real-time within live cells.