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Supplementary MaterialsS1 Fig: Examination of the neutralizing activity of mAbs against DENV4. analyzed by flow cytometry. Substitution of W212 (W210 in DENV3) led to a significant loss of binding activity of DD11-4 (A and C). Substitution of E26 led to a significant loss of binding activity of DD18-5 (B and D). Data shown are from one representative experiment of two independent Salinomycin manufacturer experiments.(TIF) pone.0136328.s002.tif (2.5M) GUID:?7A55D1E4-6735-49E4-8C40-738A5A860006 Data Availability StatementAll relevant data are within the paper Salinomycin manufacturer and its Supporting Information files. Abstract The four serotypes of dengue virus (DENV1-4) pose a serious threat to global health. Cross-reactive and non-neutralizing antibodies enhance viral infection, thereby exacerbating the disease via antibody-dependent enhancement (ADE). Studying the epitopes targeted by these enhancing antibodies would improve the immune responses against DENV infection. In order to investigate the roles Salinomycin manufacturer of antibodies in the pathogenesis of dengue, we generated a panel of 16 new monoclonal antibodies (mAbs) against DENV4. Using plaque reduction neutralization test (PRNT), we examined the neutralizing activity of these mAbs. Furthermore, we used the and ADE assay to evaluate the enhancement of DENV infection by mAbs. The results Salinomycin manufacturer indicate that the cross-reactive and poorly neutralizing mAbs, DD11-4 and DD18-5, strongly enhance DENV1-4 infection of K562 cells and increase mortality in AG129 mice. The epitope residues of these enhancing mAbs were identified using virus-like particle (VLP) mutants. W212 and E26 are the epitope residues of DD11-4 and DD18-5, respectively. In conclusion, we generated and characterized 16 new mAbs against DENV4. DD11-4 and D18-5 possessed non-neutralizing activities and enhanced viral infection. Moreover, we identified the epitope residues of enhancing mAbs on envelope protein. These results may provide useful information for development Anxa5 of safe dengue vaccine. Introduction There are an estimated 390 million dengue infections every year, mostly in tropical and subtropical areas [1]. Dengue infection can cause asymptomatic dengue fever (DF), as well as more life-threatening illness, such as dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS) [2]. Although initial infection with DENV provides immunity against the same serotype, subsequent infection by other serotypes can result in a more severe disease [3, 4]. The presence of non-neutralizing and sub-neutralizing antibodies bound to DENV exacerbates the disease by binding to the Fc receptors (FcR) of cells. This hypothetical process is termed antibody-dependent enhancement (ADE) [3, 5]. At the time of writing, there is no approved vaccine or therapy that can alleviate the symptoms of dengue infection [6]. DENV, which consists of four closely related serotypes (DENV1-4), is a member of the genus within the family [7]. The genome of DENV is a positive-strand RNA of about 11 kb in length. The viral RNA is translated into a single polyprotein that is cleaved by cellular and viral proteases into three structural proteins [capsid (C), premembrane (prM), and envelope (E) proteins] and seven non-structural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5 proteins) [8, 9]. After replication, the virus is assembled and subsequently transported to the Golgi. In the acidic environment of the trans-Golgi network (TGN), the prM protein is cleaved by furin to generate mature virions [10]. Co-expression of prM and E proteins can produce recombinant virus-like particles (VLPs), which are similar in structure and antigenicity to infectious virus particles, and have been used broadly in epitope mapping, diagnosis, and development of vaccines [11, 12]. In addition, NS1 protein, the secreted nonstructural glycoprotein, also plays Salinomycin manufacturer a critical role in pathogenesis of DENV infection. Antibodies against NS1 can bind to endothelial cells and cause apoptosis [13, 14]. The E protein is required for viral attachment to cell surface receptor(s), fusion with endosomal membranes, and entry into target cells. Thus, the E protein is regarded as an important target for neutralizing DENV [15C18]. In the mature virion, the.