fa Enzymology and Protein Chemistry Enzymology and Protein Chemistry مقاله کوتاه Short Communication <p><strong>Background:</strong> 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. <strong>Methods:</strong> 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 Ellman&#39;s test and SDS-PAGE analysis were used to evaluate free thiols produced and confirm the reduction process, respectively. <strong>Results:</strong> 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&deg;C for 30 minutes resulted in approximately 0.4, 1.2, 5.4, 7, 8, 8, and 8 thiols per antibody, respectively. <strong>Conclusion:</strong> Optimized site-specific conjugation can provide better process control and reproducibility for the development of disulfide-based ADCs.</p> Conjugation, Dithiothreitol, Antibody drug conjugate 270 274 http://ibj.pasteur.ac.ir/browse.php?a_code=A-10-1-653&slc_lang=fa&sid=1 Meghdad Abdollahpour-Alitappeh Yes Hybridoma Lab., Department OF Immunology, Pasteur Institute of Iran, Tehran, Iran Majid Lotfinia No Department of Biochemistry, Pasteur Institute of Iran, Tehran, Iran Sepand Razavi-Vakhshourpour No Hybridoma Lab., Department OF Immunology, Pasteur Institute of Iran, Tehran, Iran Saeed Jahandideh No Department of Biochemistry, Pasteur Institute of Iran, Tehran, Iran Hamid Najminejad No Hybridoma Lab., Department OF Immunology, Pasteur Institute of Iran, Tehran, Iran Koushan Sineh Sepehr No Department of Immunology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran Reza Moazami No Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran; Elnaz Shams No Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran; Mahdi Habibi-Anbouhi No National Cell Bank of Iran, Pasteur Institute of Iran, Tehran, Iran Mohsen Abolhassani No Hybridoma Lab., Department OF Immunology, Pasteur Institute of Iran, Tehran, Iran