Concerning the potential contribution of protein or peptide-display systems to study epitopes with relevant immunological features, the archaeal RAD display system stands out as a highly stable scaffold protein that allows the presentation of constrained target peptides. Here, we employed the RAD display system to present peptides derived from the SARS-CoV-2 Spike protein (S) as a tool to detect specific serum antibodies and to generate polyclonal antibodies capable of inhibit the virus infectivity in vitro. Initially, 44 selected linear S-derived peptides were genetically fused with the RAD scaffold (RAD-SCoV-epitopes), which were subsequently screened for antigenicity with sera collected from COVID-19-infected patients. In a second step, selected RAD-SCoV-epitopes were employed to immunize mice and generate antibodies. Based on phenotypic screening techniques, some of these antibodies were able to recognize replicating viral particles in VERO CCL-81. Notably, seven selected RAD-SCoV-epitopes induced antibodies capable of neutralizing viral infection. Our findings highlight the RAD display system as an useful platform for the immunological characterization of peptides and might be a valuable strategy for the design of antigens aiming peptide-based vaccines, epitope-specific antibody mapping, and development of antibodies for diagnostic and therapeutic purposes.