ArticleAnalytical chemistry2015
Method for Sorting and Pairwise Selection of Nanobodies for the Development of Highly Sensitive Sandwich Immunoassays.
Article in Analytical chemistry, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 18 papers.
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Who cites it
18 citing papers in PubMed, 45 citations in OpenAlex.
- Strategies for the Immobilization and Signal Amplification of a Double Nanobody Sandwich ELISA for Human Microsomal Epoxide Hydrolase.Analytical chemistry · 2024Article
- Mix-and-Read Nanobody-Based Sandwich Homogeneous Split-Luciferase Assay for the Rapid Detection of Human Soluble Epoxide Hydrolase.Analytical chemistry · 2023Article
- Nanobody-Based Blocking of Binding ELISA for the Detection of Anti-NS1 Zika-Virus-Specific Antibodies in Convalescent Patients.Tropical medicine and infectious disease · 2023Article
- Biophysical and biochemical insights in the design of immunoassays.Biochimica et biophysica acta. General subjects · 2023Review
- Isolation and Characterization of Single-Domain Antibodies from Immune Phage Display Libraries.Methods in molecular biology (Clifton, N.J.) · 2023Article
- A highly sensitive nanobody-based immunoassay detecting SARS-CoV-2 nucleocapsid protein using all-recombinant reagents.Frontiers in immunology · 2023Article
- Arsenal of nanobodies shows broad-spectrum neutralization against SARS-CoV-2 variants of concern in vitro and in vivo in hamster models.Communications biology · 2022Article
- Nanobody-based label-free photoelectrochemical immunoassay for highly sensitive detection of SARS-CoV-2 spike protein.Analytica chimica acta · 2022Article
- Generation of Thermally Stable Affinity Pairs for Sensitive, Specific Immunoassays.Methods in molecular biology (Clifton, N.J.) · 2022Article
- A guide to: generation and design of nanobodies.The FEBS journal · 2021Review
- Highly Sensitive Detection of Zika Virus Nonstructural Protein 1 in Serum Samples by a Two-Site Nanobody ELISA.Biomolecules · 2020Article
- Nanobody-based electrochemical competitive immunosensor for the detection of AFBMikrochimica acta · 2020Article
- Development of Improved Double-Nanobody Sandwich ELISAs for Human Soluble Epoxide Hydrolase Detection in Peripheral Blood Mononuclear Cells of Diabetic Patients and the Prefrontal Cortex of Multiple Sclerosis Patients.Analytical chemistry · 2020Article
- Development of a Simple Pretreatment Immunoassay Based on an Organic Solvent-Tolerant Nanobody for the Detection of Carbofuran in Vegetable and Fruit Samples.Biomolecules · 2019Article
- Nanobody-based binding assay for the discovery of potent inhibitors of CFTR inhibitory factor (Cif).Analytica chimica acta · 2019Article
- Nanobody Based Immunoassay for Human Soluble Epoxide Hydrolase Detection Using Polymeric Horseradish Peroxidase (PolyHRP) for Signal Enhancement: The Rediscovery of PolyHRP?Analytical chemistry · 2017Article
- Single-Domain Antibodies As Versatile Affinity Reagents for Analytical and Diagnostic Applications.Frontiers in immunology · 2017Review
- VHH antibodies: emerging reagents for the analysis of environmental chemicals.Analytical and bioanalytical chemistry · 2016Review
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Authors and funding
11 authors at 2 institutions in 1 country.
Funding
Abstract
Single domain heavychain binders (nanobodies) obtained from camelid antibody libraries hold a great promise for immunoassay development. However, there is no simple method to select the most valuable nanobodies from the crowd of positive clones obtained after the initial screening. In this paper, we describe a novel nanobody-based platform that allows comparison of the reactivity of hundreds of clones with the labeled antigen, and identifies the best nanobody pairs for two-site immunoassay development. The output clones are biotinylated in vivo in 96-well culture blocks and then used to saturate the biotin binding capacity of avidin coated wells. This standardizes the amount of captured antibody allowing their sorting by ranking their reactivity with the labeled antigen. Using human soluble epoxide hydrolase (sEH) as a model antigen, we were able to classify 96 clones in four families and confirm this classification by sequencing. This provided a criterion to select a restricted panel of five capturing antibodies and to test each of them against the rest of the 96 clones. The method constitutes a powerful tool for epitope binning, and in our case allowed development of a sandwich ELISA for sEH with a detection limit of 63 pg/mL and four log dynamic range, which performed with excellent recovery in different tissue extracts. This strategy provides a systematic way to test nanobody pairwise combinations and would have a broad utility for the development of highly sensitive sandwich immunoassays.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.