ISSN 2410-7751 (Print)
ISSN 2410-776X (Online)
"Biotechnologia Acta" v. 7, no 1, 2014
Р. 54-59, Bibliography 14, Russian
Universal Decimal classification: 577.27:578.81+[616.931:615.279]
https://doi.org/10.15407/biotech7.01.054
Oliinyk O. S., Kaberniuk A. A., Kolibo D. V., Komisarenko S. V.
Palladin Institute of Biochemistry of National Academy of Sciences of Ukraine, Kyiv, Ukraine
Diphtheria toxin is an exoantigen of Corynebacterium diphtheriae that inhibits protein synthesis and kills sensitive cells. The aim of this study was to obtain human recombinant single-chain variable fragment (scFv) antibodies against receptor-binding B subunit of diphtheria toxin.
12 specific clones were selected after three rounds of a phage display naїve (unimmunized) human antibody library against recombinant B-subunit. scFv DNA inserts from these 12 clones were digested with MvaI, and 6 unique restriction patterns were found. Single-chain antibodies were expressed in Escherichia coli XL1-blue. The recombinant proteins were characterized by immunoblotting of bacterial extracts and detection with an anti-E-tag antibody. The toxin B-subunit-binding function of the single-chain antibody was shown by ELISA. The affinity constants for different clones were found to be from 106 to 108 М–1.
Due to the fact, that these antibody fragments recognized epitopes in the receptor-binding Bsubunit of diphtheria toxin, further studies are interesting to evaluate their toxin neutralization properties and potential for therapeutic applications. Obtained scFv-antibodies can also be used for detection and investigation of biological properties of diphtheria toxin.
Key words: scFv-antibodies, diphtheria toxin, phage display, naive human recombinant antibody library.
© Palladin Institute of Biochemistry of National Academy of Sciences of Ukraine, 2014
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