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Goose parvovirus (GPV) can cause a highly contagious and fatal gosling plague (GP) disease in goslings and muscoy ducklings. Here, three goose-origin neutralizing single chain variable fragment (scFv) antibodies against GPV SYG-61 were isolated. The genes of scFv antibodies were derived from goslings immunized with GPV SYG-61, and scFvs were subcloned into a pBSD vector for the construction of pBSD-scFv libraries. The pBSD-scFv libraries were screened following three rounds using VP2 (protective antigen of GPV) as the bait by flow cytometry (FCM). After screening, the 15 clones with high mean fluorescence intensity (MFI) were isolated and sequenced. These 15 scFvs were expressed by pET-28a (+) in E. coli. The specificity and affinity of the 15 purified scFvs were successfully confirmed by ELISA. In the preliminary neutralization experiment on primary goose embryo fibroblast (GEF) in vitro, three of the 15 purified scFvs (named scFv-10, scFv-11 and scFv-50) showed significant neutralizing capacities. The study generated the first goose-origin neutralizing scFv against GPV and laid the foundation for the appearance of full-length goose-origin neutralizing monoclonal antibody against GPV.