Maebashi Institute of Technology, Gunma, 371-0816, Japan.
Department of Environmental Biology and Chemistry, Graduate School of Science and Engineering for Research, University of Toyama, Toyama, 930-8555, Japan.
Anal Chim Acta. 2017 Mar 15;958:30-37. doi: 10.1016/j.aca.2016.12.028. Epub 2016 Dec 28.
Electrochemical sensing of ovalbumin (OVA) was performed using magnetic beads with OVA recognition (RNRCKGTDVQAW)/electron-transfer (YYYYC) peptides. The focus of this study was to construct a highly sensitive and regenerative tool for OVA detection based on the interaction between a protein and peptide-1(RNRCKGTDVQAWYYYYC). The peptide-1 was introduced to the bead through four types of cross-linking reagents. Magnetic beads of different sizes with N-(6-maleimidocaproyloxy)sulfosuccinimide (Sulfo-EMCS) were also prepared. An oxidation peak due to tyrosine residues at 0.65 V depended on the distance of the electron-transfer peptide from the bead surface and on the surface area of the magnetic beads that contacted the electrode surface. The response of the electro-transfer peptide moiety was decreased because the protein was accumulated via the recognition peptide on the beads. When using Sulfo-EMCS and beads that were 6.0-6.9 μm in diameter, the calibration curve of OVA was linear and ranged from 8.0 × 10 to 2.0 × 10 M. To regenerate the magnetic beads, the measurements were achieved after removal of the OVA using a denaturing reagent. When OVA was added to fetal bovine serum containing a complex matrix, OVA was recovered at a rate of 98-100%. Consequently, these magnetic beads could be a powerful tool for the sensing of OVA in real samples.
使用具有 OVA 识别（RNRCKGTDVQAW）/电子转移（YYYYC）肽的磁性珠进行卵清蛋白（OVA）的电化学检测。本研究的重点是基于蛋白质和肽-1（RNRCKGTDVQAWYYYYC）之间的相互作用，构建一种用于 OVA 检测的高灵敏度和可再生工具。肽-1 通过四种类型的交联试剂引入到珠子上。还制备了具有 N-（6-马来酰亚胺基己酰氧基）琥珀酰亚胺（Sulfo-EMCS）的不同大小的磁性珠。在 0.65 V 处由于酪氨酸残基引起的氧化峰取决于电子转移肽距珠表面的距离以及与电极表面接触的磁性珠的表面积。由于通过珠上的识别肽累积了蛋白质，因此电转移肽部分的响应降低。当使用 Sulfo-EMCS 和直径为 6.0-6.9 μm 的珠子时，OVA 的校准曲线呈线性，范围从 8.0×10到 2.0×10 M。为了再生磁性珠，使用变性试剂去除 OVA 后进行测量。当在含有复杂基质的胎牛血清中添加 OVA 时，OVA 的回收率为 98-100％。因此，这些磁性珠可以成为用于检测实际样品中 OVA 的有力工具。
