Screening and identification of human ZnT8-specific single-chain variable fragment (scFv) from type 1 diabetes phage display library

Abstract

Zinc transporter 8 (ZnT8) is a major autoantigen and a predictive marker in type 1 diabetes (T1D). To investigate ZnT8-specific antibodies, a phage display library from T1D was constructed and single-chain antibodies against ZnT8 were screened and identified. Human T1D single-chain variable fragment (scFv) phage display library consists of approximately 110 8 clones. After four rounds of bio-panning, seven unique clones were positive by phage ELISA. Among them, C27 and C22, which demonstrated the highest affinity to ZnT8, were expressed in Escherichia coli Top10F' and then purified by affinity chromatography. C27 and C22 specifically bound ZnT8 N/C fusion protein and ZnT8 C terminal dimer with one Arg325Trp mutation. The specificity to human islet cells of these scFvs were further confirmed by immunohistochemistry. In conclusion, we have successfully constructed a T1D phage display antibody library and identified two ZnT8-specific scFv clones, C27 and C22. These ZnT8-specific scFvs are potential agents in immunodiagnostic and immunotherapy of T1D.Zinc transporter 8 (ZnT8), phage display, single-chain variable fragment (scFv), type 1 diabetes (T1D) Citation:Wu, Q., Wang, X., Gu, Y., Zhang, X., Qin, Y., Chen, H., Xu, X., Yang, T., and Zhang, M. (2016). Screening and identification of human ZnT8-specific single-chain variable fragment (scFv) from type 1 diabetes phage display library. Sci China Life Sci 59, 686 -693.

Phage Display Technology in Biomarker Identification with Emphasis on Non-Cancerous Diseases

“…Recently, autoantibodies against ZnT8 (zinc transporter protein 8), which is released by pancreatic beta islets, have been studied as a potential indicator of type 1 diabetes [82]. The scFv phage display library was used to isolate ZnT8-specific antibodies that exhibited exceptional specificity for beta cells, making them suitable for immunodiagnostic purposes [83].…”

Vaccine for Diabetes—Where Do We Stand?

“…Apart from IA-2, the zinc transporter (ZnT8) has been also studied as a major autoantigen target in T1DM immunotherapy. However, no investigations have been conducted to test ZnT8 as a diabetes vaccine in trials [ 85 , 86 , 87 , 88 ].…”

Clinical implications of phage display technique: Digging deeper in diagnostic and therapeutic grounds.

“…Zinc transporter protein 8 (ZnT8) is secreted by pancreatic beta islets and autoantibodies of this proteins have been recently studied as type 1 diabetes marker [28]. Using scFv phage display library, ZnT8-specific antibodies have been extracted which had great specificity for beta cells, having immunodiagnostic applications [29]. PACAP (pituitary adenylate cyclase activating polypeptide) is a neuropeptide that is known for its protective effects in various pathologies including diabetes [30].…”