Measuring Encapsulation Efficiency in Cell-Mimicking Giant Unilamellar Vesicles
Pashiini Supramaniam1, Zibo Wang2,3, Stelios Chatzimichail2
1Department of Chemistry, Imperial College London, London W12 0BZ, U.K.
ACS Synthetic Biology
|March 28, 2023
Summary
We developed a new method to count molecules inside artificial cells. This quality control protocol precisely measures encapsulation efficiency per vesicle, crucial for synthetic biology applications.
Area of Science:
- Synthetic biology
- Biotechnology
- Biophysics
Background:
- Bottom-up synthetic biology aims to create programmable artificial cells.
- Existing methods for generating giant unilamellar vesicle-based artificial cells are numerous.
- Quantitative measurement of molecular constituents in newly formed artificial cells remains underdeveloped.
Purpose of the Study:
- To develop and validate an artificial cell quality control (AC/QC) protocol.
- To enable absolute quantification of encapsulated biomolecules in single vesicles.
- To assess the variability of encapsulation efficiency in artificial cells.
Main Methods:
- Utilized a microfluidic-based single-molecule approach for analysis.
- Developed an artificial cell quality control (AC/QC) protocol.
- Quantified encapsulated biomolecules on a per-vesicle basis.
Main Results:
- Achieved absolute quantification of encapsulated biomolecules.
- Determined per-vesicle encapsulation efficiencies, revealing significant variability (2.4% to 41%).
- Demonstrated that desired internal concentrations can be achieved by adjusting initial concentrations, despite variability.
Conclusions:
- The developed AC/QC protocol provides precise measurement of artificial cell contents.
- Significant variability in encapsulation efficiency necessitates caution when using artificial cells as models or standards.
- This method is vital for advancing the development of functional artificial cells in synthetic biology.
