Wageningen Food Safety Research, Wageningen University and Research, 6700 AE Wageningen, The Netherlands.
Laboratory of Organic Chemistry, Wageningen University, Stippeneng 4, 6708 WE Wageningen, The Netherlands.
Biosensors (Basel). 2022 Feb 22;12(3):138. doi: 10.3390/bios12030138.
The use of peptides in immunoassays can be favored over the use of the full protein when more cost effective or less toxic approaches are needed, or when access to the full protein is lacking. Due to restricted access to recombinant bovine somatotropin (rbST), a protein enhancing growth and lactating performances of livestock, which use has been banned in the EU, Canada and Australia (amongst others), we developed a peptide-based biorecognition assay on an imaging planar array analyzer. For this, we identified the rbST epitope that is responsible for binding to the rbST-targeting monoclonal antibody 4H12 (MAb 4H12) to be DLEEGILALMR. This linear peptide was synthesized and coupled to microspheres, after which it was tested in a biorecognition competitive inhibition assay format. We observed IC values of approximately 0.11 μg mL, which are lower than observed for the full rbST protein (IC = 0.20 μg mL). Importantly, there was no binding with the scrambled peptide. Preliminary results of directly coupled peptides in a microsphere biorecognition assay for detection of rbST are presented. Real-life applicability for detection of somatotropins (STs) in injection preparations of bovine-, porcine- and equine ST are shown. This newly developed immunoassay strongly supports future developments of peptide-based immunoassays to circumvent the limited access to the full protein.
当需要更经济有效或毒性更低的方法，或者无法获得完整蛋白质时，肽在免疫测定中的应用可以优于使用完整蛋白质。由于受到限制，无法获得重组牛生长激素（rbST），这是一种增强牲畜生长和泌乳性能的蛋白质，其使用已在欧盟、加拿大和澳大利亚（等）禁止，我们在成像平面阵列分析仪上开发了基于肽的生物识别测定法。为此，我们确定了负责与针对 rbST 的单克隆抗体 4H12（MAb 4H12）结合的 rbST 表位为 DLEEGILALMR。该线性肽被合成并偶联到微球上，然后在生物识别竞争抑制测定法中进行测试。我们观察到的 IC 值约为 0.11 μg mL，低于全 rbST 蛋白的观察值（IC = 0.20 μg mL）。重要的是，与乱序肽没有结合。本文介绍了直接偶联在微球生物识别测定中检测 rbST 的肽的初步结果。展示了用于检测牛、猪和马 ST 注射制剂中生长激素（STs）的实际应用。这种新开发的免疫测定法强烈支持未来基于肽的免疫测定法的发展，以规避对完整蛋白质的有限获取。
