In Vitro Assays for Endolysin Efficacy and Safety Profiling.

This chapter describes three complementary in vitro assays designed to evaluate the antimicrobial potential and cytotoxic safety of endolysins. The Kirby-Bauer disk diffusion method enables rapid screening of endolysin activity based on inhibition zones against bacterial lawns. A serial dilution assay quantifies bactericidal efficacy through colony-forming unit (CFU) reduction, allowing dose-dependent assessment of killing activity. To assess host safety, a hemolytic assay using human red blood cells measures potential cytotoxicity by monitoring membrane lysis. Together, these methods offer a practical and reproducible framework for early-stage functional screening of candidate endolysins under controlled laboratory conditions.