Figure 2.

Characterization assays for testing activity or specificity of antimicrobial compounds. (a) Commonly used assays for measuring antimicrobial peptide (AMP) activity against individual species or strains in monoculture. (i) Broth dilution. Serial dilutions of an antimicrobial compound in liquid media are inoculated with the target bacteria. (ii) Agar diffusion. Varying doses of the antimicrobial compound are placed on agar plates inoculated with a bacterial lawn. Antimicrobial activity is measured by the zone of clearance. (iii) Metabolic activity test. A colorimetric or fluorogenic assay is used to assess metabolic activity of the target bacteria by measuring enzyme reduction of a substrate. (iv) Time-kill assay. An assay determines antimicrobial activity over time to measure rate of killing by the test compound. (b) Anaerobic culture systems designed to simulate microbiota within the human body, used to assay the AMP’s effect on microbial diversity. (i) Anaerobic batch cell culture. Small volume bioreactors are used to cultivate microbiota samples. (ii) Droplet-based cell culture. Droplet emulsions are generated using a microfluidic device to segregate a mixed microbial community and isolate low-abundance species.