An innovative approach in the detection of Toxocara canis excretory/secretory antigens using specific nanobodies

Francisco Javier Morales-Yanez, Idalia Sariego, Cecile Vincke, Gholamreza Hassanzadeh-Ghassabeh, Katja Polman, Serge Muyldermans

    Research output: Contribution to journalA1: Web of Science-articlepeer-review

    Abstract

    Human toxocariasis is a zoonosis resulting from the migration of larval stages of the dog parasite Toxocara canis into the human paratenic host. Despite its well-known limitations, serology remains the most important tool to diagnose the disease. Our objective was to employ camelid single domain antibody fragments also known as nanobodies (Nbs) for a specific and sensitive detection of Toxocara canis excretory/secretory (TES) antigens. From an alpaca immune Nb library, we retrieved different Nbs with specificity for TES antigens. Based on ELISA experiments, these Nbs did not show any cross-reactivity with Ascaris lumbricoides, Ascaris suum, Pseudoterranova decipiens, Anisakis simplex and Angiostrongylus cantonensis larval antigens. Western blot and immunocapturing revealed that Nbs 1TCE39, 1TCE52 and 2TCE49 recognise shared epitopes on different components of TES antigen. The presence of disulphide bonds in the target antigen seems to be essential for recognition of the epitopes by these three Nbs. Three separate sandwich ELISA formats, using monovalent and bivalent Nbs, were assessed to maximise the detection of TES antigens in solution. The combination of biotinylated, bivalent Nb 2TCE49 on a strep-tavidin pre-coated plate to capture TES antigens, and Nb 1TCE39 chemically coupled to horseradish peroxidase for detection of the captured TES antigens, yielded the most sensitive ELISA with a limit of detection of 0.650 ng/ml of TES antigen, spiked in serum. Moreover, the assay was able to detect TES antigens in sera from mice, taken 3 days after the animals were experimentally infected with T. canis. The specific characteristics of Nbs make this ELISA not only a promising tool for the detection of TES antigens in clinical samples, but also for a detailed structural and functional study of TES antigens. (C) 2019 The Authors. Published by Elsevier Ltd on behalf of Australian Society for Parasitology.

    Original languageEnglish
    JournalInternational Journal for Parasitology
    Volume49
    Issue number8
    Pages (from-to)635-645
    Number of pages11
    ISSN0020-7519
    DOIs
    Publication statusPublished - 2019

    Keywords

    • Toxocara canis
    • Excretory-secretory antigens
    • Single domain antibodies
    • Nanobody
    • Sandwich ELISA
    • SINGLE-DOMAIN ANTIBODIES
    • CARBOHYDRATE EPITOPES
    • LABORATORY DIAGNOSIS
    • INFECTIVE LARVAE
    • MOLECULAR-BASIS
    • IN-VITRO
    • SERODIAGNOSIS
    • PROTEIN
    • SURFACE
    • ELISA

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