The development of effective immunotherapies for solid tumors remains a significant challenge. In previous studies, we identified PVT1, a long non-coding RNA, with the peptide HF10 derived from PVT1, presented by HLA-A24. This study aims to develop a single-chain variable fragment (scFv) that specifically recognizes the HLA-A24/HF10 complex (HF10 scFv) and to evaluate its specificity, reactivity, and therapeutic potential as part of a T cell engaging bispecific antibody (HF10xCD3) in vitro and in vivo. Using a scFv phage display library, we screened for scFv clones targeting the HLA-A24/HF10 peptide complex. The selected HF10 scFv was engineered into an IgG1 format (HF10-hIgG1), which demonstrated high affinity (KD = 2.18 × 10⁻⁸ M) and specific detection of the HLA-A24/HF10 complex on HLA-A24( +)/PVT1( +) tumor cell lines. Furthermore, HF10 scFv was incorporated into a T cell engaging bispecific antibody (HF10xCD3), which induced cytotoxicity in these tumor cell lines. In a mouse xenograft model, HF10xCD3 administration exhibited significant anti-tumor activity. In conclusion, HF10xCD3 represents a promising candidate for immunotherapy targeting solid tumors.