Characterization of carbohydrate specificity of monoclonal antibodies to fungal antigenic markers using biotinylated oligosaccharides as coating antigens

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Abstract

Mannan and β-(1→3)-glucan are two polysaccharide markers that are characteristic for a number of fungal pathogens, including Candida albicans, which is the most common cause of invasive mycoses in humans. In this study, we examined the epitope specificity of two monoclonal antibodies, CM532 and FG70, which recognize certain oligosaccharide fragments from these fungal polysaccharides. Using a panel of biotinylated oligosaccharides as coating antigens, we found that antibody CM532, obtained by immunization with a pentamannoside β-Man-(1→2)-β-Man-(1→2)-α-Man-(1→2)-α-Man-(1→2)-α-Man KLH conjugate, selectively recognizes the trisaccharide β-Man-(1→2)-α-Man-(1→2)-α-Man epitope. Another antibody, FG70, obtained by immunization with heptaglucan β-Glc-(1→3)-[β-Glc-(1→3)]5-β-Glc with KLH, interacts with a linear β-(1→3)-linked pentaglucoside fragment, and the presence of 3,6-branches within this epitope does not significantly affect the interaction efficiency. The data obtained indicate that the monoclonal antibodies under consideration can be used to create currently lacking effective diagnostics for the detection of fungal infections.

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About the authors

M. L. Gening

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: v_krylov@ioc.ac.ru

Laboratory of Synthetic Glycovaccines

Russian Federation, 119991 Moscow

A. V. Polyanskaya

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: v_krylov@ioc.ac.ru

Laboratory of Synthetic Glycovaccines

Russian Federation, 119991 Moscow

A. N. Kuznetsov

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: v_krylov@ioc.ac.ru

Laboratory of Synthetic Glycovaccines

Russian Federation, 119991 Moscow

A. D. Titova

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: v_krylov@ioc.ac.ru

Laboratory of Synthetic Glycovaccines

Russian Federation, 119991 Moscow

V. I. Yudin

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: v_krylov@ioc.ac.ru

Laboratory of Synthetic Glycovaccines

Russian Federation, 119991 Moscow

D. V. Yashunskiy

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: nen@ioc.ac.ru

Laboratory of Glycoconjugate Chemistry

Russian Federation, 119991 Moscow

Y. E. Tsvetkov

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: nen@ioc.ac.ru

Laboratory of Glycoconjugate Chemistry

Russian Federation, 119991 Moscow

O. N. Yudina

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: nen@ioc.ac.ru

Laboratory of Glycoconjugate Chemistry

Russian Federation, 119991 Moscow

V. B. Krylov

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Author for correspondence.
Email: v_krylov@ioc.ac.ru

Laboratory of Synthetic Glycovaccines

Russian Federation, 119991 Moscow

N. E. Nifantiev

N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences

Email: nen@ioc.ac.ru

Laboratory of Glycoconjugate Chemistry

Russian Federation, 119991 Moscow

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Supplementary files

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1. JATS XML
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2. Scheme 1. Synthesis of 1-KLH (a) and 2-KLH (b) conjugates used as immunogens in the production of monoclonal antibodies

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3. Scheme 2. Synthesis of coating antigens for ELISA: conjugates with PAA (a) and biotin (b)

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4. Fig. 1. Results of carbohydrate specificity screening of the CM532 antibody using carbohydrate ligands 1m–14m (a), related to Candida albicans mannan. Schematic representation of immobilization of conjugates with PAA and biotin on the plate surface (b). ELISA results obtained using conjugates with polyacrylamide (c) and biotin (d); PAA – polyacrylamide

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5. Fig. 2. Results of carbohydrate specificity screening of the FG70 antibody using carbohydrate ligands 1g–13g (a), related to the common fungal marker β-(1→3)-D-glucan. b – ELISA results obtained using biotinylated conjugates

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