InVivoMAb anti-Dengue virus type 2 E protein DIII

Catalog #BE0432
Clone:
DV2-96
Reactivities:
Virus

$164.00 - $4,280.00

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Product Details

The DV2-96 monoclonal antibody reacts with sites along the lateral ridge of the DIII domain on the E protein of dengue virus serotype 2 (DENV-2). Dengue virus (DENV) is a single-stranded RNA virus that causes dengue fever and is transmitted to humans by infected female mosquitoes (Aedes aegypti). DENV infections have emerged as a global health problem in the tropics and subtropics, risking almost 50% of the world’s population. DENV has four genetically related but antigenically distinct serotypes, namely DENV-1, DENV-2, DENV-3, and DENV-4. After infection, the DENV virus particles stay viable and replicate within RBCs, platelets, and hemopoietic cells. According to scientific research, depending on pre-existing antibodies and the following infecting serotype, a DENV infection can either protect against or increase the severity of a subsequent dengue infection, e.g., DENV-2. Experimental studies have shown that a larger neutralizing antibody titer is required to protect against symptomatic DENV-2 than other serotypes, and a wide range of pre-existing anti-DENV-binding antibody titers can exacerbate DENV-2 symptoms and sickness. All DENV serotypes are neurotropic, but DENV-2 causes severe to potentially fatal deterioration of the CNS, and DENV-2 is the most frequent cause of dengue epidemics worldwide. Experimental research involving BHK21 cell-based PRNT assays demonstrated the in vitro neutralizing activity of the DV2-96 monoclonal antibody against multiple DENV-2 genotypes, such as 16681, Southeast Asian; NGC, Southeast Asian; C0477, Southeast Asian; ArA6894, Indian; IQT2913, American; and PM33974, West African. Notably, in antibody-dependent enhancement (ADE) assays using P388D1 cells, in vitro incubation of a range of concentrations of DV2-96 monoclonal antibody with immature DENV (prior to infection) promoted the infectivity of immature DENV particles through its ADE effects. In prophylactic experiments with BALB/c mice, a single in vivo dose of the DV2-96 monoclonal antibody (500 μg/mouse) one day prior to an intracranial infection with DENV-2 NGC provided significant protection against the lethal infection of DENV-2 NGC. Post-infection therapeutic experiments also demonstrated the in vivo protective efficacy of the DV2-96 monoclonal antibody against DENV-2 NGC infection in mice.

Specifications

Isotype Mouse IgG2c, κ
Recommended Dilution Buffer InVivoPure pH 7.0 Dilution Buffer
Immunogen 1:1 mixture of DENV-2 strains 16681 and NGC
Reported Applications in vivo neutralization of DENV-2
in vitro neutralization DENV-2
ELISA
Flow cytometry
Plaque reduction neutralization tests (PRNT)
Antibody-dependent enhancement (ADE)
Formulation PBS, pH 7.0
Contains no stabilizers or preservatives
Endotoxin <2EU/mg (<0.002EU/μg)
Determined by LAL gel clotting assay
Purity >95%
Determined by SDS-PAGE
Sterility 0.2 µm filtration
Production Purified from cell culture supernatant in an animal-free facility
Purification Protein G
Molecular Weight 150 kDa
Storage The antibody solution should be stored at the stock concentration at 4°C. Do not freeze.
Antibody-dependent enhancement (ADE)
Georgiev GI, Malonis RJ, Wirchnianski AS, Wessel AW, Jung HS, Cahill SM, Nyakatura EK, Vergnolle O, Dowd KA, Cowburn D, Pierson TC, Diamond MS, Lai JR. (2022). "Resurfaced ZIKV EDIII nanoparticle immunogens elicit neutralizing and protective responses in vivo" Cell Chem Biol 10.1016/j.chembiol.2022.02.004. PubMed

Zika virus (ZIKV) is a flavivirus that can cause severe disease, but there are no approved treatments or vaccines. A complication for flavivirus vaccine development is the potential of immunogens to enhance infection via antibody-dependent enhancement (ADE), a process mediated by poorly neutralizing and cross-reactive antibodies. Thus, there is a great need to develop immunogens that minimize the potential to elicit enhancing antibodies. Here we utilized structure-based protein engineering to develop "resurfaced" (rs) ZIKV immunogens based on E glycoprotein domain III (ZDIIIs), in which epitopes bound by variably neutralizing antibodies were masked by combinatorial mutagenesis. We identified one resurfaced ZDIII immunogen (rsZDIII-2.39) that elicited a protective but immune-focused response. Compared to wild type ZDIII, immunization with resurfaced rsZDIII-2.39 protein nanoparticles produced fewer numbers of ZIKV EDIII antigen-reactive B cells and elicited serum that had a lower magnitude of induced ADE against dengue virus serotype 1 (DENV1) Our findings enhance our understanding of the structural and functional determinants of antibody protection against ZIKV.

ELISA
Rockstroh A, Barzon L, Pacenti M, Palù G, Niedrig M, Ulbert S. (2015). "Recombinant Envelope-Proteins with Mutations in the Conserved Fusion Loop Allow Specific Serological Diagnosis of Dengue-Infections" PLoS Negl Trop Dis 10.1371/journal.pntd.0004218. PubMed

Dengue virus (DENV) is a mosquito-borne flavivirus and a major international public health concern in many tropical and sub-tropical areas worldwide. DENV is divided into four major serotypes, and infection with one serotype leads to immunity against the same, but not the other serotypes. The specific diagnosis of DENV-infections via antibody-detection is problematic due to the high degree of cross-reactivity displayed by antibodies against related flaviviruses, such as West Nile virus (WNV), Yellow Fever virus (YFV) or Tick-borne encephalitis virus (TBEV). Especially in areas where several flaviviruses co-circulate or in the context of vaccination e.g. against YFV or TBEV, this severely complicates diagnosis and surveillance. Most flavivirus cross-reactive antibodies are produced against the highly conserved fusion loop (FL) domain in the viral envelope (E) protein. We generated insect-cell derived recombinant E-proteins of the four DENV-serotypes which contain point mutations in the FL domain. By using specific mixtures of these mutant antigens, cross-reactivity against heterologous flaviviruses was strongly reduced, enabling sensitive and specific diagnosis of the DENV-infected serum samples in IgG and IgM-measurements. These results have indications for the development of serological DENV-tests with improved specificity.

Antibody-dependent enhancement (ADE)
De Burghgraeve T, Kaptein SJ, Ayala-Nunez NV, Mondotte JA, Pastorino B, Printsevskaya SS, de Lamballerie X, Jacobs M, Preobrazhenskaya M, Gamarnik AV, Smit JM, Neyts J. (2012). "An analogue of the antibiotic teicoplanin prevents flavivirus entry in vitro" PLoS One 10.1371/journal.pone.0037244. PubMed

There is an urgent need for potent inhibitors of dengue virus (DENV) replication for the treatment and/or prophylaxis of infections with this virus. We here report on an aglycon analogue of the antibiotic teicoplanin (code name LCTA-949) that inhibits DENV-induced cytopathic effect (CPE) in a dose-dependent manner. Virus infection was completely inhibited at concentrations that had no adverse effect on the host cells. These findings were corroborated by quantification of viral RNA levels in culture supernatant. Antiviral activity was also observed against other flaviviruses such as the yellow fever virus and the tick-borne encephalitis virus (TBEV). In particular, potent antiviral activity was observed against TBEV. Time-of-drug-addition experiments indicated that LCTA-949 inhibits an early stage in the DENV replication cycle; however, a virucidal effect was excluded. This observation was corroborated by the fact that LCTA-949 lacks activity on DENV subgenomic replicon (that does not encode structural proteins) replication. Using a microsopy-based binding and fusion assay employing DiD-labeled viruses, it was shown that LCTA-949 targets the early stage (binding/entry) of the infection. Moreover, LCTA-949 efficiently inhibits infectivity of DENV particles pre-opsonized with antibodies, thus potentially also inhibiting antibody-dependent enhancement (ADE). In conclusion, LCTA-949 exerts in vitro activity against several flaviviruses and does so (as shown for DENV) by interfering with an early step in the viral replication cycle.

Antibody-dependent enhancement (ADE), ELISA
da Silva Voorham JM, Rodenhuis-Zybert IA, Ayala Nuñez NV, Colpitts TM, van der Ende-Metselaar H, Fikrig E, Diamond MS, Wilschut J, Smit JM. (2012). "Antibodies against the envelope glycoprotein promote infectivity of immature dengue virus serotype 2" PLoS One 10.1371/journal.pone.0029957. PubMed

Cross-reactive dengue virus (DENV) antibodies directed against the envelope (E) and precursor membrane (prM) proteins are believed to contribute to the development of severe dengue disease by facilitating antibody-dependent enhancement of infection. We and others recently demonstrated that anti-prM antibodies render essentially non-infectious immature DENV infectious in Fcγ-receptor-expressing cells. Immature DENV particles are abundantly present in standard (st) virus preparations due to inefficient processing of prM to M during virus maturation. Structural analysis has revealed that the E protein is exposed in immature particles and this prompted us to investigate whether antibodies to E render immature particles infectious. To this end, we analyzed the enhancing properties of 27 anti-E antibodies directed against distinct structural domains. Of these, 23 bound to immature particles, and 15 enhanced infectivity of immature DENV in a furin-dependent manner. The significance of these findings was subsequently tested in vivo using the well-established West Nile virus (WNV) mouse model. Remarkably, mice injected with immature WNV opsonized with anti-E mAbs or immune serum produced a lethal infection in a dose-dependent manner, whereas in the absence of antibody immature WNV virions caused no morbidity or mortality. Furthermore, enhancement infection studies with standard (st) DENV preparations opsonized with anti-E mAbs in the presence or absence of furin inhibitor revealed that prM-containing particles present within st virus preparations contribute to antibody-dependent enhancement of infection. Taken together, our results support the notion that antibodies against the structural proteins prM and E both can promote pathogenesis by enhancing infectivity of prM-containing immature and partially mature flavivirus particles.

in vivo neutralization of DENV-2, in vitro neutralization DENV-2, Flow Cytometry, Plaque reduction neutralization tests (PRNT)
Sukupolvi-Petty S, Austin SK, Engle M, Brien JD, Dowd KA, Williams KL, Johnson S, Rico-Hesse R, Harris E, Pierson TC, Fremont DH, Diamond MS. (2010). "Structure and function analysis of therapeutic monoclonal antibodies against dengue virus type 2" J Virol 10.1128/JVI.01087-10. PubMed

Dengue virus (DENV) is the most prevalent insect-transmitted viral disease in humans globally, and currently no specific therapy or vaccine is available. Protection against DENV and other related flaviviruses is associated with the development of antibodies against the viral envelope (E) protein. Although prior studies have characterized the neutralizing activity of monoclonal antibodies (MAbs) against DENV type 2 (DENV-2), none have compared simultaneously the inhibitory activity against a genetically diverse range of strains in vitro, the protective capacity in animals, and the localization of epitopes. Here, with the goal of identifying MAbs that can serve as postexposure therapy, we investigated in detail the functional activity of a large panel of new anti-DENV-2 mouse MAbs. Binding sites were mapped by yeast surface display and neutralization escape, cell culture inhibition assays were performed with homologous and heterologous strains, and prophylactic and therapeutic activity was evaluated with two mouse models. Protective MAbs localized to epitopes on the lateral ridge of domain I (DI), the dimer interface, lateral ridge, and fusion loop of DII, and the lateral ridge, C-C' loop, and A strand of DIII. Several MAbs inefficiently inhibited at least one DENV-2 strain of a distinct genotype, suggesting that recognition of neutralizing epitopes varies with strain diversity. Moreover, antibody potency generally correlated with a narrowed genotype and serotype specificity. Five MAbs functioned efficiently as postexposure therapy when administered as a single dose, even 3 days after intracranial infection of BALB/c mice. Overall, these studies define the structural and functional complexity of antibodies against DENV-2 with protective potential.