Monoclonal antibodies are a branch of biological drugs that are produced in different cellular systems using recombinant DNA technology. Antibodies are glycoproteins and they are very important in the antibody immune response. During the production of large and complex proteins, post-translational modifications (PTM) are essential for their biological activity. Among the most important PTMs are glycosylation, where the attachment of sugars to the amino acid residues of the antibody takes place. Glycosylation is a very critical modification of therapeutic proteins known to change efficacy, solubility, stability (especially prior to proteolysis), immunogenicity and the rate of clearance of the active substance from the body. For this reason, the thorough characterization of the sugar residues content, the sugar chain structure and glycosylation sites that are present in the antibody is essential to ensure the safety and efficacy of the drug. Due to the complexity and obscurity of previous analytical methods, it is useful to find alternative techniques. Because glycan structures are inside the antibodies, they need to be pre-treated, which can be done in different ways. In addition to the enzymatic method where the glycans are split, thermal pretreatment is also possible, in which partial opening of the structure occurs. The primary goal of the experimental work was to find the conditions of thermal pretreatment in which there would be no aggregation. Likewise, we wanted to find a method that would confirm the opening of the molecular structure and the position where the dispersion occurs.
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