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Target-Gated Ratiometric pH Sensing via Tetrahedral DNA Framework-Based Dual-CRISPR System.

Zheng Fang1, Yaya Hao2, Xiaolei Zuo1,2

  • 1Institute of Molecular Medicine, Shanghai Key Laboratory for Nucleic Acids Chemistry and Nanomedicine, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200127, China.

Analytical Chemistry
|March 20, 2026
PubMed
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This summary is machine-generated.

Researchers developed a novel pH sensor using DNA nanotechnology and CRISPR systems. This self-calibrating nanosensor enables precise, target-activated pH detection at cell membranes, advancing the study of extracellular vesicle communication.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Nanotechnology

Background:

  • Extracellular vesicle (EV) communication is pH-dependent, influencing biogenesis, cargo release, and membrane fusion.
  • Accurate pH sensing is vital for understanding EV function and microenvironmental regulation in diseases.

Purpose of the Study:

  • To develop a self-calibrating nanosensor for ratiometric pH detection at lipid membranes.
  • To enable target-activated pH sensing for probing EV-mediated communication.

Main Methods:

  • Utilized a tetrahedral DNA framework (TDF) to coassemble orthogonal Cas12a and Cas13a CRISPR nucleases.
  • Exploited distinct pH-activity profiles of Cas12a and Cas13a for self-calibration.
  • Integrated fluorogenic reporters and programmed conditional activation via disease biomarkers like miR-146a.

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Main Results:

  • Demonstrated quantitative assembly fidelity and robust pH responsiveness across physiological ranges.
  • Achieved target-gated pH monitoring on cell-derived exosomes and during liposome fusion.
  • Validated the system's stability and reduced variability compared to conventional probes.

Conclusions:

  • Established a versatile platform for programmable, ratiometric sensing at biomembrane interfaces.
  • Offers new avenues for investigating EV communication and microenvironmental dynamics.
  • Provides a tool for disease biomarker detection and cellular process monitoring.