High-Resolution LC-MS Characterization of Antibody Drug Conjugates with PNGase F Enzymatic Cleavage of N-linked Oligosaccharides using Biozen Native-RP-5

Antibody drug conjugates (ADCs) are a promising biotherapeutic modality due to their ability to target tumor cells while sparing healthy ones. This study demonstrates the use of Biozen Native RP-5 for intact mass measurement and separation of ADC drug-to-antibody ratio (DAR) species under native reversed-phase conditions. Four cysteine-linked ADCs (Adcetris, Padcev, Polivy, and Tivdak) were enzymatically deglycosylated using PNGase F. The treated samples showed a loss of mass corresponding to N-linked glycans, confirming successful deglycosylation. The Biozen Native RP-5 column provides a clearer mass fingerprint and simplifies ADC analysis without dissociating antibody subunits.