Enzyme-Linked Immunosorbent Assay for Detection of Filovirus Species-Specific Antibodies

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The envelope glycoprotein (GP) of Marburg virus (MARV) and Ebola virus (EBOV) is responsible for virus entry into host cellsand is known as the only target of neutralizing antibodies. While knowledge about EBOV-neutralizing antibodies and the mechanism for the neutralization of infectivity is being accumulated gradually, little is known about antibodies that can efficiently regulate MARV infectivity. Here we show that MARV GP-specific monoclonal antibodies AGP127-8 (IgG1) and MGP72-17 (IgM), which do not inhibit the GP-mediated entry of MARV into host cells, drastically reduced the budding and release of progeny viruses from infected cells. These antibodies similarly inhibited the formation of virus-like particles (VLPs) consisting of GP, the viral matrix protein, and nucleoprotein, whereas the Fab fragment of AGP127-8 showed no inhibitory effect. Morphological analyses revealed that filamentous VLPs were bunched on the surface of VLP-producing cells cultured in the presence of the antibodies. These results demonstrate a novel mechanism of the antibody-mediated inhibition of MARV budding, in which antibodies arrest unformed virus particles on the cell surface. Our data lead to the idea that such antibodies, like classical neutralizing antibodies, contribute to protective immunity against MARV and that the "classical" neutralizing activity is not the only indicator of a protective antibody that may be available for prophylactic and therapeutic use.

Section: Methodsmentioning

confidence: 99%

Inhibition of Marburg Virus Budding by Nonneutralizing Antibodies to the Envelope Glycoprotein

Kajihara

Marzi

Nakayama

et al. 2012

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“…The His-EBOV-GP capture antigen was generated using truncated EBOV glycoproteins (strain Zaire) as previously described (37) Flow cytometry analysis. The frequency of CD4-or CD8-positive cells producing IFN-␥ was assessed by flow cytometry.…”

Section: Enzyme-linked Immunosorbent Assay (Elisa)mentioning

confidence: 99%

Airway Delivery of an Adenovirus-Based Ebola Virus Vaccine Bypasses Existing Immunity to Homologous Adenovirus in Nonhuman Primates

Richardson

Pillet

Bello

et al. 2013

b Anti-adenovirus serotype 5 antibodies are capable of neutralizing adenovirus serotype 5-based vaccines. In mice and guinea pigs, intranasal delivery of adenovirus serotype 5-based vaccine bypasses induced adenovirus serotype 5 preexisting immunity, resulting in protection against species-adapted Ebola virus challenge. In this study, nonhuman primates were vaccinated with adenovirus serotype 5-based vaccine either intramuscularly or via the airway route (intranasally/intratracheally) in the presence or absence of adenovirus serotype 5 preexisting immunity. Immune responses were evaluated to determine the effect of both the vaccine delivery route and preexisting immunity before and after a lethal Ebola virus (Zaïre strain Kikwit 95) challenge. Intramuscular vaccination fully protected nonhuman primates in the absence of preexisting immunity, whereas the presence of preexisting immunity abrogated vaccine efficacy and resulted in complete mortality. In contrast, the presence of preexisting immunity to adenovirus serotype 5 did not alter the survival rate of nonhuman primates receiving the adenovirus serotype 5-based Ebola virus vaccine in the airway. This study shows that airway vaccination with adenovirus serotype 5-based Ebola virus vaccine can efficiently bypass preexisting immunity to adenovirus serotype 5 and induce protective immune responses, albeit at lower efficacy than that using an intramuscular vaccine delivery route.

“…Here, we show that LLOV GP has the potential to mediate viral entry Animal Care and Use Committee. The GP-based enzyme-linked immunosorbent assay (ELISA) was performed as described previously (25). Serum samples were serially diluted with PBS containing 0.05% Tween 20, 0.5% bovine serum albumin, and 2% FCS.…”

mentioning

confidence: 99%

Characterization of the Envelope Glycoprotein of a Novel Filovirus, Lloviu Virus

Maruyama

Miyamoto

Kajihara

et al. 2014

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f Lloviu virus (LLOV), a novel filovirus detected in bats, is phylogenetically distinct from viruses in the genera Ebolavirus and Marburgvirus in the family Filoviridae. While filoviruses are known to cause severe hemorrhagic fever in humans and/or nonhuman primates, LLOV is biologically uncharacterized, since infectious LLOV has never been isolated. To examine the properties of LLOV, we characterized its envelope glycoprotein (GP), which likely plays a key role in viral tropism and pathogenicity. We first found that LLOV GP principally has the same primary structure as the other filovirus GPs. Similar to the other filoviruses, virus-like particles (VLPs) produced by transient expression of LLOV GP, matrix protein, and nucleoprotein in 293T cells had densely arrayed GP spikes on a filamentous particle. Mouse antiserum to LLOV VLP was barely cross-reactive to viruses of the other genera, indicating that LLOV is serologically distinct from the other known filoviruses. For functional study of LLOV GP, we utilized a vesicular stomatitis virus (VSV) pseudotype system and found that LLOV GP requires low endosomal pH and cathepsin L, and that human C-type lectins act as attachment factors for LLOV entry into cells. Interestingly, LLOV GP-pseudotyped VSV infected particular bat cell lines more efficiently than viruses bearing other filovirus GPs. These results suggest that LLOV GP mediates cellular entry in a manner similar to that of the other filoviruses while showing preferential tropism for some bat cells.