Volume 21, Issue 4 (7-2017)                   ibj 2017, 21(4): 270-274 | Back to browse issues page

PMID: 28410549
PMCID: PMC5459942

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Background: Reduction/alkylation is one of the leading strategies for the development of antibody drug conjugates (ADCs). Precise control of the reduction process would not only yield a defined number of free thiols per antibody but also result in development of more homogenous conjugates. Methods: In the present study, we investigated the effect of various dithiothreitol (DTT) concentrations, temperature conditions, and DTT exposure times on antibody reduction. After antibody reduction, the Ellmanchr('39')s test and SDS-PAGE analysis were used to evaluate free thiols produced and confirm the reduction process, respectively. Results: DTT concentration seems to be a potential factor in the reduction process. Concentrations of 0.1, 1, 5, 10, 20, 50, and 100 mM DTT at 37°C for 30 minutes resulted in approximately 0.4, 1.2, 5.4, 7, 8, 8, and 8 thiols per antibody, respectively. Conclusion: Optimized site-specific conjugation can provide better process control and reproducibility for the development of disulfide-based ADCs.