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Unlocking Zeptomolar Single-Molecule Detection by Synergizing Digital Microfluidics and Digital CRISPR
Zheng Li1,2,3, Fenggang Li1,4, Liyan Hua5
1Shenzhen Key Laboratory of Smart Healthcare Engineering, Guangdong Provincial Key Laboratory of Advanced Biomaterials, Department of Biomedical Engineering, Southern University of Science and Technology, Shenzhen, Guangdong 518055, China.
Journal of the American Chemical Society
|November 13, 2025
Summary
We developed a Dual-Digital immunoassay (DDA) platform for highly sensitive biomarker detection. This automated system achieves zeptomolar sensitivity, improving early disease diagnosis.
Area of Science:
- Biomarker Discovery
- Assay Development
- Molecular Diagnostics
Background:
- Accurate disease diagnosis requires sensitive, quantitative detection of immune biomarkers.
- Current methods struggle with sensitivity, specificity, and background noise.
- Ultrasensitive detection is crucial for early disease identification.
Purpose of the Study:
- Introduce the Dual-Digital immunoassay (DDA) platform.
- Demonstrate its capability for highly sensitive, automated biomarker quantification.
- Validate DDA for early disease diagnosis using clinically relevant biomarkers.
Main Methods:
- Developed a fully automated DDA platform integrating digital microfluidics and CRISPR-Cas13a amplification.
- Utilized magnetic bead-based immunocapture and digital microwell arrays.
- Optimized the assay for reduced background noise and enhanced sensitivity.
Main Results:
- Achieved zeptomolar (zM) detection limits for protein biomarkers (LOD down to 100 zM).
- Demonstrated >100-fold improvement over existing ultrasensitive assays.
- Quantified heart failure biomarkers (NT-proBNP, IL-6, TNF-α) in serum at attomolar (aM) levels.
Conclusions:
- The DDA platform offers unprecedented sensitivity and automation for biomarker analysis.
- Enables precise quantification of low-abundance biomarkers for early disease detection.
- Shows significant potential for automated screening and risk assessment across various diseases.
