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Updated: May 10, 2026

Synthesis and Bioconjugation of Thiol-Reactive Reagents for the Creation of Site-Selectively Modified Immunoconjugates
Published on: March 6, 2019
Antibody conjugation via one and two C-terminal selenocysteines
Xiuling Li1, Jiahui Yang2, Christoph Rader3
1Department of Cancer Biology, The Scripps Research Institute, Scripps Florida, Jupiter, FL 33458, USA.
Abstract:
Conventional antibody conjugation methods generate antibody-drug conjugates that are heterogeneous mixtures with undefined stoichiometry and variable pharmacokinetic and pharmacodynamic properties. We have previously described a strategy to generate site-specific antibody conjugates by genetic engineering of an antibody with a single C-terminal selenocysteine, the 21st natural amino acid, which displays unique chemical reactivity allowing selective conjugation in the presence of all other natural amino acids. In the present work, we describe a method for expanding this technology to higher drug-to-antibody ratios by genetically engineering an antibody with two C-terminal selenocysteines. Both selenocysteines effectively conjugate to a fluorescent iodoacetamide derivative and the resulting conjugate fully retains its antigen binding capability. Our method provides a platform for creating stoichiometrically defined antibody-drug conjugates for therapeutic intervention.
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