Nanobody-based immunosensor detection enhanced by photocatalytic-electrochemical redox cycling

Stanislav Trashin, Francisco Morales-Yánez, Saranya Thiruvottriyur Shanmugam, Linda Paredis, Erik N Carrión, Idalia Sariego, Serge Muyldermans, Katja Polman, Sergiu M Gorun, Karolien De Wael

Research output: Contribution to journalA1: Web of Science-article

Abstract

Detection of antigenic biomarkers present in trace amounts is of crucial importance for medical diagnosis. A parasitic disease, human toxocariasis, lacks an adequate diagnostic method despite its worldwide occurrence. The currently used serology tests may stay positive even years after a possibly unnoticed infection, whereas the direct detection of a re-infection or a still active infection remains a diagnostic challenge due to the low concentration of circulating parasitic antigens. We report a time-efficient sandwich immunosensor using small recombinant single-domain antibodies (nanobodies) derived from camelid heavy-chain antibodies specific to Toxocara canis antigens. An enhanced sensitivity to pg/mL levels is achieved by using a redox cycle consisting of a photocatalytic oxidation and electrochemical reduction steps. The photocatalytic oxidation is achieved by a photosensitizer generating singlet oxygen (1O2) that, in turn, readily reacts with p-nitrophenol enzymatically produced under alkaline conditions. The photooxidation produces benzoquinone that is electrochemically reduced to hydroquinone, generating an amperometric response. The light-driven process could be easily separated from the background, thus making amperometric detection more reliable. The proposed method for detection of the toxocariasis antigen marker shows superior performances compared to other detection schemes with the same nanobodies and outperforms by at least two orders of magnitude the assays based on regular antibodies, thus suggesting new opportunities for electrochemical immunoassays of challenging low levels of antigens.

Original languageEnglish
JournalAnalytical Chemistry
Volume93
Issue number40
Pages (from-to)13606-13614
Number of pages9
ISSN0003-2700
DOIs
Publication statusPublished - 2021

Keywords

  • Animals
  • Biosensing Techniques
  • Electrochemical Techniques
  • Humans
  • Immunoassay
  • Limit of Detection
  • Oxidation-Reduction
  • Toxocara canis
  • Toxocariasis

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