In this study, we tried to produce an ostrich-derived phage display library for capturing the best single-chain variable fragment (scFv) binders to target antigens. To this aim, two ostriches were immunized with Beta-Tc3 cells which include target antigens (PTPRN). After evaluation of the immunization process by ELISA, PMBC cells were isolated from ostrich blood. cDNA was synthesized following total RNA extraction. Variable segments of antibody (light and heavy) were amplified by PCR and the products were cloned to the PSEX81 phagemids. The resulting phagemids were electroporated into the TG1 bacteria and infected by M13KO7 helper phages to produce the phage display library. In the final steps, the released phages were introduced to the biopanning process for enrichment and screening for capturing the best binders to target antigens.