Production and Characterization of F(Ab’)2 Fragments Obtained by Enzymatic Digestion from Murine Anti-MRSA PBP2a Monoclonal Antibodies

Abstract

Infections caused by methicillin-resistant Staphylococcus aureus (MRSA) are a worldwide health problem. In a previous study, a murine monoclonal antibody (mMAB), capable of binding to PBP2a within MRSA strains, was generated. F(ab’)₂ antibody fragments are widely described in the literature as immunochemical tools and reagents for diagnostics and therapeutics, particularly because of their low immunogenicity and rapid pharmacokinetics. In this study, F(ab’)₂ fragments from mMAB were generated by enzymatic digestion, using pepsin. They were purified by affinity chromatography using protein A and concentrated by a MWCO 50 kDa filtration unit. The results indicate that it is possible to obtain F(ab’)₂ fragments by pepsin digestion. ELISA, western blotting, and fluorescence microscopy data demonstrated that F(ab’)₂ affinity for PBP2a is not lost even after the enzymatic digestion process. As expected, in the pharmacokinetics tests, F(ab’)₂ presented a faster elimination (between 12 and 18 h) compared to IgG. These F(ab’)₂ fragments could be used in future immunodiagnostic applications, including in vitro or in situ radiolabeling and in the treatment of infections caused by this important pathogen.