Adenovirus Hexon Antigen ELISA Kit

Name: Adenovirus Hexon Antigen ELISA Kit
SKU: DEIAFY103
Rating: 5 (1 reviews)

Regulatory status: For research use only, not for use in diagnostic procedures.

COA

SDS

Datasheet

Summary

Sample

Feces

Detection Method

sELISA

Intended Use

The Adenovirus ELISA is an in vitro procedure for the qualitative determination of adenovirus antigen in feces.

Contents of Kit

1. Adenovirus Antigen Capture Plate (96 tests) - 2 ea.
2. Sample Preparation Reagent (1x) - 12 ml
3. Adenovirus Positive Control (1x) - 1 ml
4. Adenovirus Negative Control (1x) - 2 x 1.5 ml
5. Wash Buffer (20x) - 2 x 50 ml
6. Adenovirus Detection Antibody, Biotin-labeled (1x) - 22 ml
7. Streptavidin-HRP (1x) - 22 ml
8. Chromagen Solution (1x) - 22 ml
9. Stop Solution (1x) - 22 ml
10. Sample Dilution Tray - 2 ea.

Storage

Store all kit components at 2-8°C. Crystal formation may occur in the wash buffer concentrate during prolonged storage at 2-8°C. The crystals can be re-dissolved by swirling the bottle in warm tap water.

General Description

Acute diarrheal disease in young children is a major cause of morbidity worldwide and is a leading cause of mortality in developing countries. Research has shown that enteric adenoviruses, primarily Ad40 and Ad41, are a leading cause of diarrhea in many of these children, second only to the rotaviruses.^(1,3,5-8) These viral pathogens have been isolated throughout the world, and can cause diarrhea in children year round.^(1-4) Infections are most frequently seen in children under two years of age,^(1-3) but have been found in patients of all ages⁽²⁾. Further studies indicate that adenoviruses are associated with 4 - 15% of all hospitalized cases of viral gastroenteritis.^(1-8)
Many laboratories use electron microscopy (EM) to detect viruses associated with gastroenteritis.^{(5, 7, 8)} Other techniques include direct genome profiling and nucleic acid hybridization, neither of which is rapid or specific.⁽⁶⁾ Alternatively, ELISA tests using Ad-specific antibodies have been shown to be a sensitive,⁽⁹⁾ specific, and rapid diagnostic method for the determination of enteric adenoviruses.⁽⁶⁾

Citations

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Background

The adenoviruses (AVs) comprise a large family of double stranded DNA viruses found in mammals, amphibians, and birds which have a common genome organization and nonenveloped icosahedral capsid structure. The capsid is made up of two types of capsomeres containing three proteins: 12 vertex capsomeres composed of fiber attachment protein and its penton base, and 240 hexons. The human adenovirus family comprises at least 49 distinct serotypes, categorized into six subgroups (A through F). Each subgroup is linked to a variety of pathologies and tissue tropisms. Adenoviruses are currently best known for their potential as therapeutic vectors, highlighted by their use as SARS-CoV-2 vaccine vehicles during the COVID-19 pandemic. A significant challenge in using adenoviruses for gene therapy and vaccine applications is the host immune response to adenovirus hexon, the major protein component of the icosahedral capsid. The primary targets of type-specific antibodies are the adenovirus structural proteins hexon and fiber. While antifiber antibodies neutralize virus infectivity only in vitro and not in vivo, the neutralizing antibody response crucial for vector applications is directed against hexon.

Hexon is the largest and most abundant of the structural proteins in the icosahedral adenovirus capsid. Ribbon representation of the Ad2 hexon subunit. The view is perpendicular to the molecular threefold axis from the inside of the molecule. The top of the molecule, which contains the loops (DE1, FG1, and FG2), forms the outer surface of the viral capsid. The hexon base contains a small loop (DE2) and two eight-stranded "viral" jellyrolls (V1 and V2), which are separated by the connector, VC. The eight jellyroll β strands are labeled B to I. The N-terminal loop, NT, lies underneath the base.

The structure of Adenovirus Hexon (John J. Rux, 2003) Figure 1. Hexon structure.
(Source: John J. Rux. et al., 2003.)

Historically, adenoviruses have played pivotal roles as molecular biology tools in the discovery of eukaryotic biological mechanisms such as RNA splicing and protein folding. Adenoviruses are maintained episomally in infected cells and do not integrate with the host genome. Consequently, the transgene is lost as cells infected with a recombinant adenovirus vector turn over so that repeated administrations of vector are necessary. Problems then arise with preexisting or acquired antiadenoviral immunity. A conservative approach to overcoming this obstacle is to design and use adenovirus-based vectors with modified hexons. After decades of research, a HAdV-based gene therapy was recently approved by regulators in the United States of America. In addition, vaccines based on modified adenoviruses against Ebola, HIV, ZIKV, RSV, and SARS-CoV2 are currently in various phases of clinical trials or already being used in the clinic.

References

1. John J. Rux. et al., Structural and Phylogenetic Analysis of Adenovirus Hexons by Use of High-Resolution X-Ray Crystallographic, Molecular Modeling, and Sequence-Based Methods. J Virol. 2003 Sep;77(17):9553-66.
2. Dhillon A, et al., Structural insights into the interaction between adenovirus C5 hexon and human lactoferrin. J Virol. 2024, 98:e01576-23.

Documents

pdf COA

pdf SDS

pdf Datasheet

Reagents for Adenovirus Assay Development

QA & Reviews

Q & A

Ask a Question

Q: I’d like to know which types of Adenovirus the ELISA Kit can target?

A: Yes, the assay detects all human Adenovirus serovars.

Q: If the ELISA kit is targeting Hexon antigen from Ad5.

A: The assay detects all human Adenovirus serovars. Ad5 will be detected.

Customer Reviews

References

Structural insights into the interaction between adenovirus C5 hexon and human lactoferrin.

J Virol.

Authors: Dhillon A, Persson BD, Volkov AN, Sülzen H, Kádek A, Pompach P, Kereïche S, Lepšík M, Danskog K, Uetrecht C, Arnberg N, Zoll S.

Abstract

Adenovirus (AdV) infection of the respiratory epithelium is common but poorly understood. Human AdV species C types, such as HAdV-C5, utilize the Coxsackie-adenovirus receptor (CAR) for attachment and subsequently integrins for entry. CAR and integrins are however located deep within the tight junctions in the mucosa where they would not be easily accessible. Recently, a model for CAR-independent AdV entry was proposed. In this model, human lactoferrin (hLF), an innate immune protein, aids the viral uptake into epithelial cells by mediating interactions between the major capsid protein, hexon, and yet unknown host cellular receptor(s). However, a detailed understanding of the molecular interactions driving this mechanism is lacking. Here, we present a new cryo-EM structure of HAdV-5C hexon at high resolution alongside a hybrid structure of HAdV-5C hexon complexed with human lactoferrin (hLF). These structures reveal the molecular determinants of the interaction between hLF and HAdV-C5 hexon. hLF engages hexon primarily via its N-terminal lactoferricin (Lfcin) region, interacting with hexon's hypervariable region 1 (HVR-1). Mutational analyses pinpoint critical Lfcin contacts and also identify additional regions within hLF that critically contribute to hexon binding. Our study sheds more light on the intricate mechanism by which HAdV-C5 utilizes soluble hLF/Lfcin for cellular entry. These findings hold promise for advancing gene therapy applications and inform vaccine development.

Development of monoclonal antibodies targeting the conserved fragment of hexon protein to detect different serotypes of human adenovirus

J Virol.

Authors: Wu L, Lin Y, Yin J, Yang C, Jiang Y, Zhai L, Wang Y, Zhu L, Wu Q, Zhang B, Wan C, Zhao W, Yang Y, Shen C, Xiao W.

Abstract

Human adenovirus (HAdV) infects the respiratory system, thus posing a threat to health. However, immunodiagnostic reagents for human adenovirus are limited. This study aimed to develop efficient diagnostic reagents based on monoclonal antibodies for diagnosing various human adenovirus infections. Evolutionary and homology analyses of various human adenoviral antigen genes revealed highly conserved antigenic fragments. The prokaryotic expression system was applied to recombinant penton, hexon, and IVa2 conserved fragments of adenovirus, which were injected into BALB/c mice to prepare human adenovirus-specific monoclonal antibodies. Enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence assay (IFA), and Western blotting were used to determine the immune specificity of the monoclonal antibodies. Indirect ELISA showed that monoclonal antibodies 1F10, 8D3, 4A1, and 9B2 were specifically bound to HAdV-3 and HAdV-55 and revealed high sensitivity and low detection limits for various human adenoviruses. Western blotting showed that 1F10 and 8D3 specifically recognized various human adenovirus types, including HAdV-1, HAdV-2, HAdV-3, HAdV-4, HAdV-5, HAdV-7, HAdV-21, and HAdV-55, and 4A1 specifically recognized HAdV-1, HAdV-2, HAdV-3, HAdV-5, HAdV-7, HAdV-21, and HAdV-55. IFAs showed that 1F10, 8D3, and 4A1 exhibited highly selective localization to A549 cells infected with HAdV-3 and HAdV-55. Finally, two antibody pairs that could detect hexon antigens HAdV-3 and HAdV-55 at low concentrations were developed. The monoclonal antibodies developed in this study show potential for detecting human adenoviruses.