The human skin has microbiological, chemical, and physical barriers that help it defend itself from invading pathogens and transient microbes. Measuring the functionality of such barriers has been challenging and requires direct application of the microbes to the test sites on human subjects. We have developed a novel ex vivo assay that can be used to measure the collective functionality of the skin defense components, especially the chemical barrier that includes antimicrobial peptides (AMPs) and antimicrobial lipids (AMLs), which can be extracted using buffer scrub methods. A defined skin area is sampled using a buffer scrub technique with a low salt acidic pH buffer and the collected sample centrifuged to reduce the level of interfering microbial load and cell debris. The supernatant is used in a modified in vitro microdilution assay designed to evaluate antimicrobial activity against test microbes. The antibacterial activity of the supernatants is quantified by plating on growth media for viable bacteria. The AMP levels in the samples are measured by standard ELISA. The major benefits of this technique are its ability to measure skin’s collective antimicrobial functional activity, and it can do so safely in an in vitro assay against bacteria and viruses that normally would not be permissible for tests on human subjects. This assay could serve to enhance our understanding of the functionality of the innate skin defenses under normal (and potentially dysbiotic) conditions for different populations to understand their role in health and disease prevention. This will also significantly bridge the gap between the existing in vitro and in vivo methods.

The human skin has microbiological, chemical, and physical barriers that help it defend itself from invading pathogens and transient microbes. Measuring the functionality of such barriers has been challenging and requires direct application of the microbes to the test sites on human subjects. We have developed a novel _ex vivo_ assay that can be used to measure the collective functionality of the skin defense components, especially the chemical barrier that includes antimicrobial peptides (AMPs) and antimicrobial lipids (AMLs), which can be extracted using buffer scrub methods. A defined skin area is sampled using a buffer scrub technique with a low salt acidic pH buffer and the collected sample centrifuged to reduce the level of interfering microbial load and cell debris. The supernatant is used in a modified _in vitro_ microdilution assay designed to evaluate antimicrobial activity against test microbes. The antibacterial activity of the supernatants is quantified by plating on growth media for viable bacteria. The AMP levels in the samples are measured by standard ELISA. The major benefits of this technique are its ability to measure skin’s collective antimicrobial functional activity, and it can do so safely in an _in vitro_ assay against bacteria and viruses that normally would not be permissible for tests on human subjects. This assay could serve to enhance our understanding of the functionality of the innate skin defenses under normal (and potentially dysbiotic) conditions for different populations to understand their role in health and disease prevention. This will also significantly bridge the gap between the existing _in vitro_ and _in vivo_ methods.

ESDR202 - An ex vivo method to assay skin’s innate defense

50th Annual Meeting of the European Society for Dermatological Research. VIRTUAL MEETING