20.500.12556/RUL-181020 Persistence and Fab-specific interaction of acyl-protein thioesterase-1 with monoclonal antibodies during downstream processing Difficult-to-remove host cell proteins (HCPs) can compromise monoclonal antibody (mAb) product quality and stability. This study investigated the persistence of acyl-protein thioesterase-1, a polysorbate-degrading HCP, during downstream processing and its interaction with a set of mAbs. Acyl-protein thioesterase-1 remained in the eluates after reprocessing a specific mAb with protein A affinity liquid chromatography, even when various wash buffer additives were used, indicating a strong product:HCP interaction that resists conventional purification strategies. Upon cleaving the mAb and purifying its Fab and Fc fragments, we used biolayer interferometry to show that acyl-protein thioesterase-1 selectively and tightly binds to the Fab region. Domain-specific sensor-immobilization approach suggested that the binding site was located on the antibody's CH1 domain, and the interaction was influenced by preincubation with the mAb's cognate antigen. These findings highlight the importance of characterizing HCP:mAb interactions as a basis for improving HCP reduction strategies. monoclonal antibodies chromatography host cell proteins acyl-protein thioesterase-1 protein-protein interactions biolayer interferometry monoklonska protitelesa kromatografija proteini gostiteljskih celic acil-protein tioesteraza-1 interakcije protein-protein interferometrija biološke plasti true false true Angleški jezik Slovenski jezik Članek v reviji 2026-03-23 09:25:30 2026-03-23 09:25:36 2026-03-24 04:26:37 0000-00-00 00:00:00 2026 0 0 Str. 1-7 art. 106920 Vol. 241 Jul. 2026 0000-00-00 Zaloznikova Objavljeno NiDoloceno 0000-00-00 0000-00-00 0000-00-00 577 1046-5928 10.1016/j.pep.2026.106920 272547075 1-s2.0-S1046592826000434-main.pdf 1-s2.0-S1046592826000434-main.pdf 1 DABDA5571EA0524A018E097542953989 91259e2114e111205caa94281fd06586fbddfa225111c9d3262b7f6e108fd884 0ed7c034-2692-11f1-b0ab-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=231083 https://www.sciencedirect.com/science/article/pii/S1046592826000434 1 c23d8c17-2691-11f1-b0ab-0050569b8976 https://repozitorij.uni-lj.si/Dokument.php?lang=slv&id=231082 Fakulteta za farmacijo 0 0 0