Oliveira-Silva Rui, Wang Yuyang, Nooteboom Sjoerd W, Prazeres Duarte M F, Paulo Pedro M R, Zijlstra Peter
MBx Molecular Biosensing, Department of Applied Physics and Institute for Complex Molecular Systems, Eindhoven University of Technology, P.O. Box 513, 5600 MB Eindhoven, The Netherlands.
iBB - Institute for Biotechnology and Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, 1049-001 Lisboa, Portugal.
ACS Appl Opt Mater. 2023 Oct 4;1(10):1661-1669. doi: 10.1021/acsaom.3c00226. eCollection 2023 Oct 27.
We have established a label-free plasmonic platform that monitors proteolytic activity in real time. The sensor consists of a random array of gold nanorods that are functionalized with a design peptide that is specifically cleaved by thrombin, resulting in a blueshift of the longitudinal plasmon. By monitoring the plasmon of many individual nanorods, we determined thrombin's proteolytic activity in real time and inferred relevant kinetic parameters. Furthermore, a comparison to a kinetic model revealed that the plasmon shift is dictated by a competition between peptide cleavage and thrombin binding, which have opposing effects on the measured plasmon shift. The dynamic range of the sensor is greater than two orders of magnitude, and it is capable of detecting physiologically relevant levels of active thrombin down to 3 nM in buffered conditions. We expect these plasmon-mediated label-free sensors to open the window to a range of applications stretching from the diagnostic and characterization of bleeding disorders to fundamental proteolytic and pharmacological studies.
我们建立了一个无标记的等离子体平台，可实时监测蛋白水解活性。该传感器由金纳米棒的随机阵列组成，这些金纳米棒用一种设计肽进行功能化，该设计肽可被凝血酶特异性切割，导致纵向等离子体发生蓝移。通过监测许多单个纳米棒的等离子体，我们实时确定了凝血酶的蛋白水解活性，并推断出相关的动力学参数。此外，与动力学模型的比较表明，等离子体位移由肽切割和凝血酶结合之间的竞争决定，这对测量的等离子体位移具有相反的影响。该传感器的动态范围大于两个数量级，并且能够在缓冲条件下检测低至3 nM的生理相关水平的活性凝血酶。我们期望这些等离子体介导的无标记传感器能够为一系列应用打开窗口，从出血性疾病的诊断和表征到基础蛋白水解和药理学研究。
