Characterization and applications of Nanobodies against human procalcitonin selected from a novel naïve Nanobody phage display library

Yan, Junrong; Wang, Pingyan; Zhu, Min; Li, Guanghui; Romão, Ema; Xiong, Sheng; Wan, Yakun

doi:10.1186/s12951-015-0091-7

Cited by 46 publications

(24 citation statements)

References 28 publications

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“…The RBDs of SARS-CoV-2 spikes were used as antigens to conduct camel immunization, then the immunized phage display library was generated according to our established methods 45,46 .…”

Section: Camel Immunization and Phage Display Library Constructionmentioning

confidence: 99%

A potent neutralizing nanobody against SARS-CoV-2 with inhaled delivery potential

Gai¹,

Li³

et al. 2020

Preprint

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The outbreak of COVID-19 has emerged as a global pandemic. The unprecedented scale and severity call for rapid development of effective prophylactics or therapeutics. We here reported Nanobody (Nb) phage display libraries derived from four camels immunized with the SARS-CoV-2 spike receptor-binding domain (RBD), from which 381 Nbs were identified to recognize SARS-CoV-2-RBD. Furthermore, seven Nbs were shown to block interaction of human angiotensin converting enzyme 2 (ACE2) with SARS-CoV-2-RBD-variants, bat-SL-CoV-WIV1-RBD and SARS-CoV-1-RBD. Among the seven candidates, Nb11-59 exhibited the highest activity against authentic SARS-CoV-2 with ND50 of 0.55 μg/mL. Nb11-59 can be produced on a large-scale in Pichia pastoris, with 20 g/L titer and 99.36% purity. It also showed good stability profile, and nebulization did not impact its stability. Overall, Nb11-59 might be a promising prophylactic and therapeutic molecule against COVID-19, especially through inhalation delivery.

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“…The RBDs of SARS-CoV-2 spikes were used as antigens to conduct camel immunization, then the immunized phage display library was generated according to our established methods 45,46 .…”

Section: Camel Immunization and Phage Display Library Constructionmentioning

confidence: 99%

A potent neutralizing nanobody against SARS-CoV-2 with inhaled delivery potential

Gai¹,

Li³

et al. 2020

Preprint

Self Cite

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“…3-phenoxybenzoic acid 4 , aflatoxin 10 , ochratoxin 8 , and tetrabromobisphenol A 9,11 . However, for analysis of large analytes, a number of sandwich immunoassays using nanobodies showed moderate performance for procalcitonin 12 , epidermal growth factor receptor 13 , Bacillus thuringiensis Cry1Ac and Cry1Fa toxins 14,15 , influenza H3N2 and H5N1 16,17 , and hepatitis B surface antigen 18 , with the LOD in the ng/mL range or even much higher. Thus, it is often important to increase the sensitivity of nanobody based immunoassays.…”

Section: Introductionmentioning

confidence: 99%

Nanobody Based Immunoassay for Human Soluble Epoxide Hydrolase Detection Using Polymeric Horseradish Peroxidase (PolyHRP) for Signal Enhancement: The Rediscovery of PolyHRP?

Cui

Morisseau

et al. 2017

Anal. Chem.

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Soluble epoxide hydrolase (sEH) is a potential pharmacological target for treating hypertension, vascular inflammation, cancer, pain and multiple cardiovascular related diseases. A variable domain of the heavy chain antibody (termed sdAb, nanobody or VHH) possesses advantages of small size, high stability, ease of genetic manipulation, and ability for continuous manufacture, making such nanobody a superior choice as an immunoreagent. In this work, we developed an ultrasensitive nanobody based immunoassay for human sEH detection using polymeric horseradish peroxidase (PolyHRP) for signal enhancement. Llama nanobodies against human sEH were used as the detection antibody in sandwich ELISAs with polyclonal anti-sEH as the capture antibody. A conventional sandwich ELISA using a HRP labeled anti-HA tag as the tracer showed a marginal sensitivity (0.0015 OD•mL/ng) and limit of detection (LOD) of 3.02 ng/mL. However, the introduction of the PolyHRP as the tracer demonstrated a 141-fold increase in the sensitivity (0.21 OD•mL/ng) and 57-fold decrease in LOD (0.05 ng/mL). Systematic comparison of three different tracers in four ELISA formats demonstrated the overwhelming advantage of PolyHRP as a label for nanobody based immunoassay. This enhanced sEH immunoassay was further evaluated in terms of selectivity against other epoxide hydrolases and detection of the target protein in human tissue homogenate samples. Comparison with an enzyme activity based assay and a Western-blot for sEH detection reveals good correlation with the immunoassay. This work demonstrates increased competiveness of nanobodies for practical sEH protein detection utilizing PolyHRP. It is worthwhile to rediscover the promising potential of PolyHRP in nanobody and other affinity based methods after its low-profile existence for decades.

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“…Surface plasmon resonance imaging (SPRi) binding assays were performed in accordance with our previously reported study (Yan et al., ). Briefly, each of the Cry1B‐specific Nbs was spotted and immobilized on the NanoCapture 3D‐chip surface for 1 h. Afterward, the chip was incubated in 2 ml of 1 mol L −1 ethanolamine‐HCl (pH 8.5) for 20 min in order to block the activated NHS groups.…”

Section: Methodsmentioning

confidence: 99%

Sensitive detection of Bacillus thuringiensis Cry1B toxin based on camel single‐domain antibodies

Zhong

et al. 2018

MicrobiologyOpen

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Bt Cry1B toxin, a residue in insect‐resistant transgenic plants, has been identified to be harmful to human health. Therefore, it is urgent to detect the Cry1B toxin level in each kind of transgenic plant. Nbs, with prominently unique physiochemical properties, are becoming more and more promising tools in the detection of target antigens. In this study, an immune phage display library that was of high quality was successfully constructed for the screening of Cry1B‐specific Nbs with excellent specificity, affinity, and thermostable. Subsequently, a novel sandwich ELISA for Cry1B detection was established, which was based on the biotin–streptavidin system using these aforementioned Nbs. This established detection system presented a linear working range from 5 to 1000 ng ml−1 and a low detection limit of 3.46 ng ml−1. The recoveries from spiked samples were in the range of 82.51%–113.56% with a relative standard deviation (RSD) lower than 5.00%. Taken together, the proposed sandwich ELISA would be a potential method for the detection of Cry1B toxin in transgenic Bt plants specifically and sensitively.

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Characterization and applications of Nanobodies against human procalcitonin selected from a novel naïve Nanobody phage display library

Cited by 46 publications

References 28 publications

A potent neutralizing nanobody against SARS-CoV-2 with inhaled delivery potential

A potent neutralizing nanobody against SARS-CoV-2 with inhaled delivery potential

Nanobody Based Immunoassay for Human Soluble Epoxide Hydrolase Detection Using Polymeric Horseradish Peroxidase (PolyHRP) for Signal Enhancement: The Rediscovery of PolyHRP?

Sensitive detection of Bacillus thuringiensis Cry1B toxin based on camel single‐domain antibodies

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