In this label-free bioassay, an electrochemiluminescence (ECL) immunosensor was developed for the quantification of breast cancer using HER-2 protein as a metastatic biomarker. For this purpose, the ECL emitter, [Ru(bpy)3]2+, was embedded into biocompatible chitosan (CS) polymer. The prepared bio-composite offered high ECL reading due to the depletion of human epidermal growth factor receptor 2 (HER-2) protein. Reduced graphene oxide (rGO) was used as substrate to increase signal stability and achieve greater sensitivity. For this, rGO was initially placed electrochemically on the glassy carbon electrode (GCE) surface by cyclic voltammetry (CV) technique. Next, the prepared CS/[Ru(bpy)3]2+ biopolymer solution was coated on a drop of the modified electrode such that the amine groups of CS and the carboxylic groups of rGO could covalently interact. Using EDC/NHS chemistry, monoclonal antibodies (Abs) of HER-2 were linked to CS/[Ru(bpy)3]2+/rGO/GCE via amide bonds between the carboxylic groups of Ab molecules and amine groups of CS. The electrochemical behavior of the electrode was studied using different electrochemical techniques such as electrochemical impedance spectroscopy (EIS), differential pulse voltammetry (DPV) and square wave voltammetry (SWV) and also ECL tests. After passing all optimization steps, the lower limit of detection (LLOQ) and linear dynamic range (LDR) of HER-2 protein were practically obtained as 1 fM and 1 fM to 1 nM, individually. Importantly, the within and between laboratory precisions were performed and the suitable relative standard deviations (RSDs) were recorded as 3.1 and 3.5%, respectively. As a proof of concept, the designed immunosensor was desirably applied for the quantification of HER-2 protein in breast cancer suffering patients. As a result, the designed ECL-based immunosensor has the capability of being used as a conventional test method in biomedical laboratories for early detection of HER-2 protein in biological fluids.

中文翻译：

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使用还原型氧化石墨烯/壳聚糖作为纳米生物相容性平台对HER-2蛋白进行乳腺癌的超灵敏生物测定

在这种无标记的生物测定中，开发了一种电化学发光（ECL）免疫传感器，用于使用HER-2蛋白作为转移性生物标记物对乳腺癌进行定量。为此，将ECL发射体[Ru（bpy）3] 2+嵌入生物相容性壳聚糖（CS）聚合物中。由于人表皮生长因子受体2（HER-2）蛋白的消耗，所制备的生物复合材料提供了较高的ECL读数。还原的氧化石墨烯（rGO）用作底物以增加信号稳定性并获得更高的灵敏度。为此，首先通过循环伏安法（CV）将rGO电化学方式放置在玻碳电极（GCE）表面上。接下来，将制备的CS / [Ru（bpy）3] 2+生物聚合物溶液涂覆在一滴修饰电极上，以使CS的胺基和rGO的羧基共价相互作用。使用EDC / NHS化学方法，通过Ab分子的羧基与CS的胺基之间的酰胺键将HER-2的单克隆抗体（Abs）与CS / [Ru（bpy）3] 2 + / rGO / GCE连接。使用不同的电化学技术，例如电化学阻抗谱（EIS），差分脉冲伏安法（DPV）和方波伏安法（SWV）以及ECL测试，研究了电极的电化学行为。通过所有优化步骤后，实际上获得的HER-2蛋白的检测下限（LLOQ）和线性动态范围（LDR）分别为1 fM和1 fM至1 nM。重要的是，进行了实验室内部和实验室之间的精确度，并且合适的相对标准偏差（RSD）分别记录为3.1％和3.5％。作为概念证明，所设计的免疫传感器理想地用于定量乳腺癌患者中的HER-2蛋白。结果，设计的基于ECL的免疫传感器具有被用作生物医学实验室中常规检测方法的能力，从而可以早期检测生物流体中的HER-2蛋白。