Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System

García-García, Aina; Madrid, Raquel; Garcia-Calvo, Eduardo; Mendoza-Chamizo, Belén; Garcı́a, Teresa; Martı́n, Rosario

doi:10.3390/foods9121838

Cited by 7 publications

(6 citation statements)

References 42 publications

(23 reference statements)

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“…Besides, the ingestion of gluten-containing cereals would trigger gluten-related disorders. Garcia-Garcia A et al [32] developed a sandwich ELISA to detect gluten in food products. The yeast P. pastoris was employed to produce a recombinant single-domain antibody (dAb8E) by phage display technology, which acted as a novel biorecognition molecule to develop an immunoassay with a LOD of 0.12 ng mL -1 and a linear range of 0.1-10 µg mL -1 .…”

Section: Protein Biomarkersmentioning

confidence: 99%

Nanobody and Nanozyme‐Enabled Immunoassays with Enhanced Specificity and Sensitivity

Wang

Xianyu

2022

Small Methods

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treatment of cancers, the second leading cause of death in the world that are hard to be treated in the terminal stages. [2] For instance, liver cancer is one of the most lethal cancers and has a poor prognosis with a 5-year survival rate of less than 20%. [3] The early detection and timely treatment of cancers are very important for the patients to save their lives. Microorganism contamination in food is the main cause of foodborne diseases and 70% of patients with foodborne illnesses get sick after taking foods or water contaminated with pathogenic microorganisms. [4] Viruses are one of the major causes of infectious diseases and various public health events like the acquired immunodeficiency syndrome (AIDS), the severe acute respiratory syndrome (SARS), and the coronavirus disease 2019 (COVID-19). [5] Therefore, the timely and rapid detection of microorganisms is of great importance to ensure human health and public safety. [6] In addition, the detections of small molecules including mycotoxins and exogenous pollutions play an important role in food safety. Mycotoxins such as aflatoxin B1, ochratoxin A, and tenuazonic acid are classified as naturally occurring carcinogens by the World Health Organization that are extremely harmful to both humans and animals. [7] They not only directly threaten human life by contaminating food, but also indirectly affect human health through the food chain by contaminating feeds of animals. [8,9] Commonly used immunoassays include enzyme-linked immunosorbent assay (ELISA), quantum dot fluorescence immunoassay, fluorescence polarization immunoassay, colloidal gold immunoassay, chemiluminescent immunoassay (CLIA), electrochemical immunoassay, and microarray-based immunoassay. In spite of the different formats of immunoassays, they share the same principle that relies on the specific binding between antigenic determinants and antibody recognition sites. Therefore, it is of great significance to prepare specific antibodies with high affinity towards the target molecules to improve the specificity of immunoassays. Conventional immunoassays generally use polyclonal and monoclonal antibodies, but both types of antibodies have limitations such as poor specificity, low stability, and complicated preparation. To meet these challenges, a new type of antibody called "nanobody" has emerged. Nanobody is the heavy-chain-only antibody with the advantages of small size, high specificity, mass expression and high stability, which can act as a substitute for conventional antibodies in immunoassays and a promising low-cost immuno-affinity detection reagent.Immunoassay as a rapid and convenient method for detecting a variety of targets has attracted tremendous interest with its high specificity and sensitivity. Among the commonly used immunoassays, enzyme-linked immunosorbent assay has been widely used as a gold standard method in various fields that consists of two main components including a recognition element and an enzyme label. With the rapid advances in nanotechnology, nanobodies and nanozym...

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Section: Protein Biomarkersmentioning

confidence: 99%

Nanobody and Nanozyme‐Enabled Immunoassays with Enhanced Specificity and Sensitivity

Wang

Xianyu

2022

Small Methods

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“…Currently, the most common methods for quantitative detection of wheat gluten, include the antibody-based immunoassay method [e.g., enzyme-linked immunosorbent assay (ELISA)] and DNA-based method [e.g., quantitative real-time polymerase chain reaction (qPCR)]. García-García et al reported a sandwiched ELISA for gluten detection with a limit of detection (LOD) of 0.12 µg/mL [15].…”

Section: Introductionmentioning

confidence: 99%

A novel AgNPs/MOF substrate-based SERS sensor for high-sensitive on-site detection of wheat gluten

Fu¹,

Du²,

Zhou³

et al. 2024

Food Science and Human Wellness

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Gluten, known as the major allergen in wheat, has gained increasing concerns in industrialized countries, resulting in an urgent need for accurate, high-sensitive, and on-site detection of wheat gluten in complex food systems. Herein, we proposed an AgNPs/MOF substrate-based surface-enhanced Raman scattering (SERS) sensor for the high-sensitive on-site detection of wheat gluten. The detection occurred on the newly in-situ synthesized AgNPs/MOF-modified SERS substrate, providing an enhancement factor (EF) of 1.89×10 5 . Benefitting from the signal amplification function of AgNPs/MOF and the superiority of SERS, this sensor represented high sensitivity performance and a wide detection range from 1×10 -15 to 2×10 -6 M with a detection limit of 1.16×10 -16 M, which allowed monitoring the trace of wheat gluten in complex food system without matrix interference. This reliable sandwich SERS sensor may provide a promising platform for high-sensitive, accurate, and on-site detection of allergens in the field of food safety.

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“…Novel single-domain (dAb) recombinant antibodies [23], suitable for the development of gluten detection ELISA systems, have been recently obtained from directed evolution processes based on phage display technology [24]. In this work, the use of Fabs is proposed instead of dAbs because of their enhanced stability and a bigger paratope [25].…”

Section: Introductionmentioning

confidence: 99%

Production and Characterization of Novel Fabs Generated from Different Phage Display Libraries as Probes for Immunoassays for Gluten Detection in Food

Garcia-Calvo,

García-García,

Rodríguez

et al. 2023

Foods

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Gluten is the main fraction of wheat proteins. It is widely used in the food industry because of the properties that are generated in the dough, but it is also able to trigger diseases like allergies, autoimmunity processes (such as celiac disease), and intolerances in sensitized persons. The most effective therapy for these diseases is the total avoidance of gluten in the diet because it not only prevents damage but also enhances tissue healing. To ensure the absence of gluten in food products labeled as gluten-free, accurate detection systems, like immunoassays, are required. In this work, four recombinant Fab antibody fragments, selected by phage display technology, were produced and tested for specificity and accuracy against gluten in experimental flour mixtures and commercial food products. A high-affinity probe (Fab-C) was identified and characterized. An indirect ELISA test was developed based on Fab-C that complied with the legal detection limits and could be applied in the assessment of gluten-free diets.

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Production of a Recombinant Single-Domain Antibody for Gluten Detection in Foods Using the Pichia pastoris Expression System

Cited by 7 publications

References 42 publications

Nanobody and Nanozyme‐Enabled Immunoassays with Enhanced Specificity and Sensitivity

Nanobody and Nanozyme‐Enabled Immunoassays with Enhanced Specificity and Sensitivity

A novel AgNPs/MOF substrate-based SERS sensor for high-sensitive on-site detection of wheat gluten

Production and Characterization of Novel Fabs Generated from Different Phage Display Libraries as Probes for Immunoassays for Gluten Detection in Food

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