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Resonant-Cantilever-Detected Kinetic/Thermodynamic Parameters for Aptamer–Ligand Binding on a Liquid–Solid Interface

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posted on 01.07.2020, 12:33 authored by Xuefeng Wang, Yarong Cheng, Shengran Cai, Jianzhong Chen, Pengcheng Xu, Ying Chen, Haitao Yu, Tiegang Xu, Sen Zhang, Xinxin Li
Nucleic acid aptamers have been widely used as recognition elements on various biosensing interfaces, but quantitative kinetic/thermodynamic analysis for revealing the aptamer–ligand binding mechanism, which occurs on a liquid–solid interface, has not been realized due to a lack of usable biophysical tools. Herein we apply a resonant microcantilever sensor to continuously record the frequency shift according to the binding-induced mass change on the liquid–solid interface. The frequency-shift curve is used for tracing the reaction process and is fitted with classic equations to calculate a set of kinetic/thermodynamic parameters, such as rate constants (ka = 902.95 M–1 s–1, kd = 0.000141 s–1), equilibrium constants (KD = 1.55 μM), the Gibbs free energy (ΔG° = −32.57 kJ/mol), and the activation energy (Ea = 38.03 kJ/mol) for the immobilized aptamer and free ATP. This quantitative analysis method is label-free, calibration-free, and highly sensitive. The kinetic/thermodynamic parameter detection method provides new resolution to the in-depth understanding of the ligand–aptamer interaction on the liquid–solid interface for biosensing or lab-on-a-chip applications.

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