Purification of chicken IgY by binding capture using elastin-like polypeptide-tagged immunoglobulin-binding domain of streptococcal protein G

Xia, Wenlong; Lu, Huipeng; Li, Yangyang; Cao, Jun; Zhou, Xiaohui; Zhang, Xinyu; Xia, Xiaoli; Sun, Huaichang

doi:10.1016/j.vetimm.2017.09.002

Cited by 10 publications

(8 citation statements)

References 18 publications

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“…The recombinant plasmid pELP-p15 was transformed into E. coli BL21 (DE3), and protein expression was induced with 0.2 mM IPTG at 20 • C for 16 h. The cells were harvested and disrupted via sonication for 10 min. Both supernatant and precipitation were collected and analyzed using 12% SDS-PAGE after centrifugation at 8000× g for 10 min at 4 • C. The ELP-p15 fusion protein was then purified using ITC, as described in our previous study [28]. First, 2 mL of bacterial lysate supernatant was incubated with 3 M (final concentration) of NaCl for 10 min at 22, 24, 26, 28, and 30 • C, respectively.…”

Section: Protein Expression and Purificationmentioning

confidence: 99%

Developing an Indirect ELISA for the Detection of African Swine Fever Virus Antibodies Using a Tag-Free p15 Protein Antigen

Wu,

Lu,

Zhu

et al. 2023

Viruses

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African swine fever (ASF) is one of the most severe diseases caused by the ASF virus (ASFV), causing massive economic losses to the global pig industry. Serological tests are important in ASF epidemiological surveillance, and more antigen targets are needed to meet market demand for ASFV antibody detection. In the present study, ASFV p15 protein was fusion-expressed in Escherichia coli (E. coli) with elastin-like polypeptide (ELP), and the ELP-p15 protein was purified using a simple inverse transition cycling (ITC) process. The ELP tag was cleaved off using tobacco etch virus protease (TEVp), resulting in a tag-free p15 protein. Western blot analysis demonstrated that the p15 protein reacted strongly with ASFV-positive serum. The p15 protein was used as a coating antigen in an indirect ELISA (iELISA) for detecting ASFV antibodies. The p15-iELISA method demonstrated high specificity to ASFV-positive sera, with a maximum detection dilution of 1:1600. Moreover, the method exhibited good reproducibility, with less intra-assay and inter-assay CV values than 10%. Therefore, p15-iELISA offers a novel approach for accurately detecting ASFV antibodies with significant clinical application potential.

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Section: Protein Expression and Purificationmentioning

confidence: 99%

Developing an Indirect ELISA for the Detection of African Swine Fever Virus Antibodies Using a Tag-Free p15 Protein Antigen

Wu,

Lu,

Zhu

et al. 2023

Viruses

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“…Some novel IgY ligands have been discovered and reported to be effective for efficient purification of IgY. − For example, Dong et al screened a synthetic ligand that had a high IgY-binding capacity (74.8 mg of IgY per mL of gel) and achieved high purity (92%) and recovery rate (78%) of egg yolk IgY. However, to date, all the identified ligands − are still not commercially available for research and development. An alternative approach is to immobilize specific target antigen to a solid phase for affinity purification of the corresponding IgY from egg yolk.…”

Section: Challenges Of Igy-based Passive Immunizationmentioning

confidence: 99%

“…The protein A or protein G affinity chromatography is widely used for efficient purification of mammalian IgG; however, such a chromatography approach does not work for IgY purification due to the lack of affinity of the Fc region of IgY to protein A or protein G . Some novel IgY ligands have been discovered and reported to be effective for efficient purification of IgY. − For example, Dong et al screened a synthetic ligand that had a high IgY-binding capacity (74.8 mg of IgY per mL of gel) and achieved high purity (92%) and recovery rate (78%) of egg yolk IgY. However, to date, all the identified ligands − are still not commercially available for research and development.…”

Section: Challenges Of Igy-based Passive Immunizationmentioning

confidence: 99%

Egg Yolk Antibody for Passive Immunization: Status, Challenges, and Prospects

Wang

Zhong

Lin

2023

J. Agric. Food Chem.

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The immunoglobulin Y (IgY) derived from hyperimmune egg yolk is a promising passive immune agent to combat microbial infections in humans and livestock. Numerous studies have been performed to develop specific egg yolk IgY for pathogen control, but with limited success. To date, the efficacy of commercial IgY products, which are all delivered through an oral route, has not been approved or endorsed by any regulatory authorities. Several challenging issues of the IgY-based passive immunization, which were not fully recognized and holistically discussed in previous publications, have impeded the development of effective egg yolk IgY products for humans and animals. This review summarizes major challenges of this technology, including in vivo stability, purification, heterologous immunogenicity, and repertoire diversity of egg yolk IgY. To tackle these challenges, potential solutions, such as encapsulation technologies to stabilize IgY, are discussed. Exploration of this technology to combat the COVID-19 pandemic is also updated in this review.

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“…Conventional affinity chromatographic methods using protein A or protein G columns cannot be performed for IgY purification, since IgYs, contrary to IgGs, do not bind to protein A or G[ 66 ]. Other types of ligands are therefore required, such as the elastin-like polypeptide-tagged immunoglobulin-binding domain of streptococcal protein G[ 67 ]. Still other ligands, such as IgY-binding peptides screened from a random peptide library, have been also proposed as a means of IgY purification[ 68 ].…”

Section: Development and Evaluation Of Egg Yolk-derived Polyclonal Igysmentioning

confidence: 99%

IgY technology: Methods for developing and evaluating avian immunoglobulins for the in vitro detection of biomolecules

Karachaliou¹,

Vassilakopoulou²,

Livaniou³

2021

WJM

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The term “IgY technology” was introduced in the literature in the mid 1990s to describe a procedure involving immunization of avian species, mainly laying hens and consequent isolation of the polyclonal IgYs from the “immune” egg yolk (thus avoiding bleeding and animal stress). IgYs have been applied to various fields of medicine and biotechnology. The present article will deal with specific aspects of IgY technology, focusing on the currently reported methods for developing, isolating, evaluating and storing polyclonal IgYs. Other topics such as current information on isolation protocols or evaluation of IgYs from different avian species are also discussed. Specific advantages of IgY technology ( e.g. , novel antibody specificities that may emerge via the avian immune system) will also be discussed. Recent in vitro applications of polyclonal egg yolk-derived IgYs to the field of disease diagnosis in human and veterinary medicine through in vitro immunodetection of target biomolecules will be presented. Moreover, ethical aspects associated with animal well-being as well as new promising approaches that are relevant to the original IgY technology ( e.g. , development of monoclonal IgYs and IgY-like antibodies through the phage display technique or in transgenic chickens) and future prospects in the area will also be mentioned.

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Purification of chicken IgY by binding capture using elastin-like polypeptide-tagged immunoglobulin-binding domain of streptococcal protein G

Cited by 10 publications

References 18 publications

Developing an Indirect ELISA for the Detection of African Swine Fever Virus Antibodies Using a Tag-Free p15 Protein Antigen

Developing an Indirect ELISA for the Detection of African Swine Fever Virus Antibodies Using a Tag-Free p15 Protein Antigen

Egg Yolk Antibody for Passive Immunization: Status, Challenges, and Prospects

IgY technology: Methods for developing and evaluating avian immunoglobulins for the in vitro detection of biomolecules

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