Comprehensive profiling of genotype-specific dengue antibody responses using a newly developed single-round infectious particle panel

Neutralizing antibodies provide critical protection against dengue virus (DENV) infections. However, under certain conditions, antibodies can also enhance infection and contribute to severe disease through antibody-dependent enhancement (ADE). ADE can occur through extrinsic mechanisms, in which antibody-opsonized virions engage with Fcγ receptors (FcγRs) on target cells to facilitate increased viral entry, while intrinsic mechanisms have been proposed in which FcγR-mediated uptake triggers intracellular pathways that promote viral replication. The extensive genetic diversity of DENV further complicates the evaluation of antibody functionality, as antibodies may exhibit distinct neutralizing and enhancing activities against different viral genotypes. Here, we developed a single-round infectious particle (SRIP) panel representing 17 DENV genotypes across all four DENV serotypes. This genotype-inclusive platform enables assessment of antibody-mediated neutralization and FcγR-dependent enhancement of infection through luciferase reporter quantification. Using this panel, we evaluated antibody responses from DENV-infected individuals, healthy volunteers, and flavivirus-cross-reactive or DENV-specific monoclonal antibodies. Our findings reveal genotype-specific variations in antibody-mediated neutralization and enhancement, emphasizing the complex interplay between antibody responses and DENV genetic diversity. This SRIP panel provides a robust approach for profiling functional antibodies against genetically diverse DENV strains. While the current platform primarily quantifies extrinsic ADE, its single-round infection format and intracellular replicon-based readout provide a foundation for future investigations of intrinsic ADE mechanisms. Overall, this platform offers broad applications for studying dengue immunity, evaluating vaccine-induced antibody responses, and advancing our understanding of antibody-mediated protection and disease enhancement. IMPORTANCE: Dengue virus (DENV) comprises four serotypes and substantial genetic diversity, which can influence the ability of antibodies to neutralize or enhance infection. However, comparative assessment of antibody functionality across diverse DENV genotypes remains challenging because differences in viral genetic backgrounds can confound functional measurements. We developed a single-round infectious particle (SRIP) panel representing 17 DENV genotypes in which the prM and E structural proteins are varied on a common replicon backbone. This design provides a standardized platform for directly comparing antibody-mediated neutralization and Fcγ receptor-dependent enhancement across genetically diverse DENV strains. By characterizing antibody responses against a broad range of genotypes, the panel can reveal differences in functional antibody activity that may be overlooked when using a limited number of representative strains. The platform is therefore useful for investigating how DENV genetic diversity influences antibody-mediated protection and extrinsic antibody-dependent enhancement (ADE), as well as for evaluating the breadth of antibody responses induced by infection or vaccination. Although the current platform primarily quantifies extrinsic ADE, its single-round infection format and quantitative intracellular reporter readout provide a foundation for future studies investigating intrinsic ADE in more physiologically relevant immune cell models. This genotype-inclusive panel offers a practical approach for standardized functional profiling of dengue antibodies and may facilitate improved evaluation of vaccine-induced immunity and antibody responses across the genetic diversity of DENV.