Measuring Thousands of Single-Vesicle Leakage Events Reveals the Mode of Action of Antimicrobial Peptides

Anal Chem. 2022 Jul 12;94(27):9530-9539. doi: 10.1021/acs.analchem.1c03564. Epub 2022 Jun 27.


Host defense or antimicrobial peptides hold promise for providing new pipelines of effective antimicrobial agents. Their activity quantified against model phospholipid membranes is fundamental to a detailed understanding of their structure-activity relationships. However, classical characterization assays often lack the ability to achieve this insight. Leveraging a highly parallelized microfluidic platform for trapping and studying thousands of giant unilamellar vesicles, we conducted quantitative long-term microscopy studies to monitor the membrane-disruptive activity of archetypal antimicrobial peptides with a high spatiotemporal resolution. We described the modes of action of these peptides via measurements of the disruption of the vesicle population under the conditions of continuous peptide dosing using a range of concentrations and related the observed modes to the molecular activity mechanisms of these peptides. The study offers an effective approach for characterizing membrane-targeting antimicrobial agents in a standardized manner and for assigning specific modes of action to the corresponding antimicrobial mechanisms.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Infective Agents* / pharmacology
  • Antimicrobial Cationic Peptides* / chemistry
  • Antimicrobial Cationic Peptides* / pharmacology
  • Antimicrobial Peptides
  • Phospholipids / chemistry
  • Unilamellar Liposomes / chemistry


  • Anti-Infective Agents
  • Antimicrobial Cationic Peptides
  • Antimicrobial Peptides
  • Phospholipids
  • Unilamellar Liposomes