Hu Jiumei, Zhang Pengfei, Shao Fangchi, Wang Tza-Huei
Department of Mechanical Engineering, Johns Hopkins University, Baltimore, MD, United States.
Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, United States.
Front Bioeng Biotechnol. 2024 Nov 21;12:1462203. doi: 10.3389/fbioe.2024.1462203. eCollection 2024.
The detection of protein biomarkers presenting at low concentrations in biological fluids is essential for disease diagnosis and therapeutic monitoring. While magnetic beads-based solid-phase immunoassays have shown promise in achieving high sensitivity for detecting low-abundance proteins, existing protocols suffer from limitations such as the cumbersome need for bead blocking and washing steps to minimize adsorption of non-specific biomolecules. These extra requirements lead to increased assay complexity and the risk of procedural errors. In this study, we present a streamlined magnetic proximity extension assay (MagPEA) using poly (oligo (ethylene glycol) methacrylate) (POEGMA)-coated beads. The polymer brush on bead surface, on the one hand, provides an effective mechanism for repelling non-specifically bound biomolecules that contribute to background signal generation without performing any bead blocking and washing steps. On the other hand, it facilitates the immobilization of capture antibodies on bead surface by simply embedding the antibodies onto the porous polymer under vacuum. Using the human inflammatory factor IL-8 as a demonstration, we show that the incorporation of POEGMA beads into MagPEA workflow significantly simplifies assay procedure while maintains high sensitivity.
检测生物体液中低浓度存在的蛋白质生物标志物对于疾病诊断和治疗监测至关重要。虽然基于磁珠的固相免疫测定法在检测低丰度蛋白质方面显示出高灵敏度的前景，但现有方案存在局限性，例如需要繁琐的磁珠封闭和洗涤步骤以尽量减少非特异性生物分子的吸附。这些额外的要求导致检测复杂性增加和操作失误的风险。在本研究中，我们提出了一种使用聚（甲基丙烯酸寡聚乙二醇酯）（POEGMA）包被磁珠的简化磁邻近延伸分析（MagPEA）方法。磁珠表面的聚合物刷一方面提供了一种有效的机制，可排斥导致背景信号产生的非特异性结合生物分子，而无需进行任何磁珠封闭和洗涤步骤。另一方面，通过在真空下将捕获抗体简单地嵌入多孔聚合物中，它有助于捕获抗体固定在磁珠表面。以人类炎症因子IL-8为例，我们表明将POEGMA磁珠纳入MagPEA工作流程可显著简化检测程序，同时保持高灵敏度。
