Digital Counting of Biomolecules Using Engineered Functional DNA Superstructures.

Area of Science:

• Biotechnology
• Molecular Biology
• Nanotechnology

Background:

• A critical need exists for simple, effective biosensing platforms for single biomolecule detection.
• Current methods face challenges in sensitivity and versatility for diverse biological, medical, and environmental applications.

Purpose of the Study:

• To develop a versatile and sensitive fluorescence counting strategy for quantifying proteins and microRNAs.
• To demonstrate the utility of functional DNA superstructures (3D DNA) as biolabels for enhanced biosensing.

Main Methods:

• Engineered highly fluorescent 3D DNA biolabels with specific recognition elements for target molecules.
• Utilized a sandwich complex formation between the 3D DNA biolabel, target, and surface-bound capture molecules.
• Employed traditional fluorescence microscopy for resolving and quantifying the formed complexes.

Main Results:

• Successfully quantified β-lactamase (bacterial hydrolase) with a detection limit of 100 aM.
• Quantified miR-21 (cancer-overexpressed microRNA) with a detection limit of 1 fM.
• Demonstrated broad utility through distinct 3D DNA biolabel designs for different targets.

Conclusions:

• The developed 3D DNA-based fluorescence counting strategy provides a versatile and sensitive platform for biomolecule quantification.
• This approach shows significant potential for applications in chemical biology, medical diagnostics, and environmental biosensing.
• The high sensitivity achieved (aM to fM range) enables detection of low-abundance targets.