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

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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.

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Fig. 1
Fig. 2

Abbreviations

EBC:

epidemic brazilian clone

ELISA:

enzyme-linked immunosorbent assay

MRSA:

methicillin-resistant Staphylococcus aureus

MSSA:

methicillin-sensitive Staphylococcus aureus

MWCO:

molecular weight cutoff

OD:

optical density

PBS:

phosphate-buffered saline

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Acknowledgements

The authors are very grateful to all technicians of the Animal Experimentation Laboratory (Fiocruz) who provided technical support for the animal experimentation assays.

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Authors and Affiliations

  1. Bio-Manguinhos, Oswaldo Cruz Foundation, Brazilian Ministry of Health, Rio de Janeiro, RJ, Brazil

    Anna Erika Vieira de Araujo, Natalia Plinio de Souza, Alvaro Paiva Braga de Sousa & Jose Procopio Moreno Senna

  2. Oswaldo Cruz Institute, Oswaldo Cruz Foundation, Brazilian Ministry of Health, Rio de Janeiro, RJ, Brazil

    Flavio Alves Lara

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  1. Anna Erika Vieira de Araujo
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  2. Natalia Plinio de Souza
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  3. Alvaro Paiva Braga de Sousa
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Correspondence to Anna Erika Vieira de Araujo.

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de Araujo, A.E.V., de Souza, N.P., de Sousa, A.P.B. et al. Production and Characterization of F(Ab’)2 Fragments Obtained by Enzymatic Digestion from Murine Anti-MRSA PBP2a Monoclonal Antibodies. Appl Biochem Biotechnol 185, 72–80 (2018). https://doi.org/10.1007/s12010-017-2624-z

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