Demystifying Low Endotoxin Recovery (LER): Poster presented at PDA Microbiology Conference 2025

Low Endotoxin Recovery (LER) is described as the inability to recover a known concentration of Control Standard Endotoxin (CSE), or Reference Standard Endotoxin (RSE) over time. It is a two-part reaction requiring a chelator and a surfactant. The chelator strips away the divalent cations causing the LPS aggregate to disassemble. Since Lipid A of LPS is hydrophobic, polysorbate encapsulates the LPS in a lipid micelle. This form of endotoxin is unable to react with Factor C of the Limulus Amebocyte Lysate.The FDA and EMA require all new biologic drugs submitted for approval to be assessed for the presence of the LER phenomenon.Organisms used to produce RSE/CSE standards are grown under high nutrient conditions including divalent cations, then purified. The divalent cations allow the organisms to develop "salt bridges" between adjacent lipopolysaccharide molecules. However, organisms in purified water systems are not exposed to divalent cations in their environment and adapt to this environment via the PhoP/PhoQ system on the bacterial surface, holding the outer membrane together. These adapted organisms' endotoxin is not affected by the chelator, and the aggregates remain intact. This is the endotoxin that will contaminate the product if a breach in the water purification system happens.