Rheumatoid arthritis (RA) is the most common chronic autoimmune disease worldwide. Although progress has been made in RA treatment in recent decades, remission cannot be effectively achieved for a considerable proportion of RA patients. Thus, novel potential targets for therapeutic strategies are needed. Semaphorin 5A (SEMA5A) plays a pivotal role in RA progression by facilitating pannus formation, and it is a promising therapeutic target. In this study, we sought to develop an antibody treatment strategy targeting SEMA5A and evaluate its therapeutic effect using a collagen-induced arthritis (CIA) model. We generated SYD12–12, a fully human SEMA5A blocking antibody, through phage display technology. SYD12–12 intervention effectively inhibited angiogenesis and aggressive phenotypes of RA synoviocytes in vitro and dose-dependently inhibited synovial hyperplasia, pannus formation, bone destruction in CIA mice. Notably, SYD12–12 also improved the Treg/Th17 imbalance in CIA mice. We confirmed through immunofluorescence and molecular docking that SYD12–12 integrated with the unique TSP-1 domain of SEMA5A. In conclusion, we developed and characterized a fully human SEMA5A-blocking antibody for the first time. SYD12–12 effectively alleviated disease progression in CIA mice by inhibiting pannus formation and improving the Treg/Th17 imbalance, demonstrating its potential for the RA treatment.