Single-Cell Caspase-3 Measurement Using a Biomimetic Nanochannel

Area of Science:

• Biomedical Engineering
• Analytical Chemistry
• Cell Biology

Background:

• Direct single-cell analysis of caspase-3 (Casp-3) is crucial for understanding disease mechanisms but remains technically challenging.
• Casp-3 plays a vital role in apoptosis and various pathological conditions.
• Existing methods often lack the sensitivity or specificity for precise single-cell quantification.

Purpose of the Study:

• To develop a novel method for direct single-cell analysis of intracellular caspase-3 (Casp-3).
• To establish a biomimetic nanochannel system for ionic detection of Casp-3 activity.
• To demonstrate the utility of this system in monitoring cellular responses to stimuli and during apoptosis.

Main Methods:

• Fabrication of a biomimetic nanochannel using a glass nanopipette.
• Immobilization of target-specific organic molecules (luc-DEVD) within the nanochannel orifice.
• Utilizing changes in ionic response due to Casp-3-mediated cleavage of luc-DEVD for detection.
• Application in probing intracellular Casp-3 fluctuations and quantifying Casp-3 during induced apoptosis.

Main Results:

• Successfully devised a biomimetic nanochannel for single-cell sampling and ionic detection of intracellular Casp-3.
• Demonstrated that Casp-3 cleavage of luc-DEVD induces detectable changes in the nanochannel's ionic properties.
• Validated the system's applicability by monitoring Casp-3 levels in response to drug stimulation and during apoptosis.
• Achieved ionic single-cell Casp-3 analysis.

Conclusions:

• The developed biomimetic nanochannel provides a viable platform for direct ionic detection of single-cell caspase-3.
• This approach offers a new perspective for single-cell protein analysis, particularly for enzymes like Casp-3.
• The technology has potential applications in studying Casp-3-related diseases and drug discovery.