In this work, rare specific binders to botulinum neurotoxin antigen were selected from human phage immunized library through three rounds of panning in vitro. Of them, clone ScFvB17 is of high affinity and specificity to target antigen. B17 is of 750bp encoding 250 amino acid. Analysis results of gene structure showed V-genes of B17 belonging respectively to VH4 or? chainⅡ family are new antibody Fv genes. More important result showed that recombinant ScFvB17 expressed in E.coli could bind specific for neurotoxin competing with antiserum. Successful selection of human phage antibody library and obtaining specific ScFv against botulinum neurotoxin serotype A provide useful materials for further study on detection of neurotoxin and human engineering therapeutic antitoxin.