Department of Chemical & Biomolecular Engineering and Electronics Design Center, Case Western Reserve University, 10900 Euclid Avenue, Cleveland, OH 44106, USA.
Dip. Chimica Materiali e Ing. Chimica "Giulio Natta", Politecnico di Milano, Via Mancinelli 7, 20131 MIlan, Italy.
Biosensors (Basel). 2017 Jul 20;7(3):29. doi: 10.3390/bios7030029.
A simple in vitro biosensor for the detection of β-amyloid 42 in phosphate-buffered saline (PBS) and undiluted human serum was fabricated and tested based on our platform sensor technology. The bio-recognition mechanism of this biosensor was based on the effect of the interaction between antibody and antigen of β-amyloid 42 to the redox couple probe of K₄Fe(CN)₆ and K₃Fe(CN)₆. Differential pulse voltammetry (DPV) served as the transduction mechanism measuring the current output derived from the redox coupling reaction. The biosensor was a three-electrode electrochemical system, and the working and counter electrodes were 50 nm thin gold film deposited by a sputtering technique. The reference electrode was a thick-film printed Ag/AgCl electrode. Laser ablation technique was used to define the size and structure of the biosensor. Cost-effective roll-to-roll manufacturing process was employed in the fabrication of the biosensor, making it simple and relatively inexpensive. Self-assembled monolayers (SAM) of 3-Mercaptopropionic acid (MPA) was employed to covalently immobilize the thiol group on the gold working electrode. A carbodiimide conjugation approach using -(3-dimethylaminopropyl)-'-ethylcarbodiimide hydrochloride (EDC) and -hydroxysuccinimide (NHS) was undertaken for cross-linking antibody of β-amyloid 42 to the carboxylic groups on one end of the MPA. The antibody concentration of β-amyloid 42 used was 18.75 µg/mL. The concentration range of β-amyloid 42 in this study was from 0.0675 µg/mL to 0.5 µg/mL for both PBS and undiluted human serum. DPV measurements showed excellent response in this antigen concentration range. Interference study of this biosensor was carried out in the presence of Tau protein antigen. Excellent specificity of this β-amyloid 42 biosensor was demonstrated without interference from other species, such as T-tau protein.
我们基于平台传感器技术，制作并测试了一种用于检测磷酸盐缓冲盐水（PBS）和未稀释人血清中β-淀粉样蛋白 42 的简单体外生物传感器。该生物传感器的生物识别机制基于β-淀粉样蛋白 42 抗体与抗原相互作用对 K₄Fe(CN)₆ 和 K₃Fe(CN)₆ 氧化还原偶探针的影响。差分脉冲伏安法（DPV）作为测量源于氧化还原偶联反应的电流输出的转换机制。生物传感器是一个三电极电化学系统，工作电极和对电极是通过溅射技术沉积的 50nm 厚金薄膜。参比电极是厚膜印刷的 Ag/AgCl 电极。激光烧蚀技术用于定义生物传感器的尺寸和结构。在生物传感器的制造中采用了具有成本效益的卷对卷制造工艺，使其简单且相对便宜。采用 3-巯基丙酸（MPA）自组装单分子层（SAM）将巯基共价固定在金工作电极上。使用 -(3-二甲基氨基丙基)-'-乙基碳化二亚胺盐酸盐（EDC）和 -羟基琥珀酰亚胺（NHS）的碳二亚胺偶联方法将β-淀粉样蛋白 42 的抗体交联到 MPA 一端的羧酸基团上。β-淀粉样蛋白 42 的抗体浓度为 18.75μg/mL。本研究中β-淀粉样蛋白 42 的浓度范围为 0.0675μg/mL 至 0.5μg/mL，适用于 PBS 和未稀释人血清。DPV 测量在该抗原浓度范围内显示出出色的响应。在 Tau 蛋白抗原存在的情况下进行了该生物传感器的干扰研究。该β-淀粉样蛋白 42 生物传感器表现出出色的特异性，没有受到其他物质（如 T-tau 蛋白）的干扰。
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