Department of Chemistry, Bio-X, and the Biophysics Program, Stanford University, Stanford, CA 94305.
Department of Materials Science and Engineering, South University of Science and Technology of China, Shenzhen 518055, China.
Proc Natl Acad Sci U S A. 2017 Sep 19;114(38):10196-10201. doi: 10.1073/pnas.1711169114. Epub 2017 Sep 5.
Identified as a major biomarker for type 1 diabetes (T1D) diagnosis, zinc transporter 8 autoantibody (ZnT8A) has shown promise for staging disease risk and disease diagnosis. However, existing assays for ZnT8 autoantibody (ZnT8A) are limited to detection by soluble domains of ZnT8, owing to difficulties in maintaining proper folding of a full-length ZnT8 protein outside its native membrane environment. Through a combined bioengineering and nanotechnology approach, we have developed a proteoliposome-based full-length ZnT8 self-antigen (full-length ZnT8 proteoliposomes; PLR-ZnT8) for efficient detection of ZnT8A on a plasmonic gold chip (pGOLD). The protective lipid matrix of proteoliposomes improved the proper folding and structural stability of full-length ZnT8, helping PLR-ZnT8 immobilized on pGOLD (PLR-ZnT8/pGOLD) achieve high-affinity capture of ZnT8A from T1D sera. Our PLR-ZnT8/pGOLD exhibited efficient ZnT8A detection for T1D diagnosis with ∼76% sensitivity and ∼97% specificity ( = 307), superior to assays based on detergent-solubilized full-length ZnT8 and the C-terminal domain of ZnT8. Multiplexed assays using pGOLD were also developed for simultaneous detection of ZnT8A, islet antigen 2 autoantibody, and glutamic acid decarboxylase autoantibody for diagnosing T1D.
锌转运体 8 自身抗体（ZnT8A）被确定为 1 型糖尿病（T1D）诊断的主要生物标志物，它在疾病风险分期和疾病诊断方面显示出了巨大的潜力。然而，由于难以在其天然膜环境之外维持全长 ZnT8 蛋白的正确折叠，现有的 ZnT8 自身抗体（ZnT8A）检测方法仅限于可溶性 ZnT8 结构域的检测。通过结合生物工程和纳米技术方法，我们开发了一种基于蛋白脂质体的全长 ZnT8 自身抗原（全长 ZnT8 蛋白脂质体；PLR-ZnT8），用于在等离子体金芯片（pGOLD）上高效检测 ZnT8A。蛋白脂质体的保护性脂质基质改善了全长 ZnT8 的正确折叠和结构稳定性，有助于固定在 pGOLD 上的 PLR-ZnT8（PLR-ZnT8/pGOLD）从 T1D 血清中高亲和力捕获 ZnT8A。我们的 PLR-ZnT8/pGOLD 用于 T1D 诊断时，对 ZnT8A 的检测具有约 76%的灵敏度和约 97%的特异性（=307），优于基于去污剂溶解的全长 ZnT8 和 ZnT8 的 C 末端结构域的检测方法。还使用 pGOLD 开发了多重检测方法，用于同时检测 ZnT8A、胰岛抗原 2 自身抗体和谷氨酸脱羧酶自身抗体，以诊断 T1D。
