Joint Department of Biomedical Engineering, North Carolina State University, Raleigh, NC, 27695-7907, USA.
Department of Materials Science and Engineering, North Carolina State University, Raleigh, NC, 27695-7907, USA.
Biosens Bioelectron. 2022 Nov 15;216:114592. doi: 10.1016/j.bios.2022.114592. Epub 2022 Aug 10.
Square wave voltammetry serves as an effective analytical means to evaluate antigen-antibody coupling at the solid-liquid interface. Herein, we describe 3-aminopropyltrimethoxysilane (APTMS) induced irreversible immobilization of anti-leptin to micellar gold nanoparticles (AuNPs). Antibodies (Abs) were orthogonally loaded on micellized AuNP assemblies via amino residual groups. The ratio of bound Ab molecules was determined by the Bradford assay. The AuNP/Ab layer modified electrodes with variable antibody surface coverage (∼400 ± 55-200 ± 30 Ab/NP) were analyzed in terms of change in backward, net current (Ip) components. The rate of antigen coupling was found to be consistent with the variation in antibody density as well as the binding affinity. The lowest detection limit was observed at the femtomolar level (0.25 fM/mL) over a wide range of antigen concentration (6.2 ng/mL to 0.12 fg/mL). The variables affecting the epitope-paratope interaction were further optimized using a chemometric approach and a response surface methodology (RSM).
方波伏安法是评估固液界面上抗原-抗体偶联的有效分析手段。本文描述了 3-氨丙基三甲氧基硅烷（APTMS）诱导的莱菔素抗体在胶束金纳米粒子（AuNPs）上的不可逆固定化。通过氨基残留基团将抗体（Abs）垂直加载到胶束化的 AuNP 组装体上。通过 Bradford 测定法确定结合的 Ab 分子的比例。根据反向、净电流（Ip）分量的变化，分析了具有可变抗体表面覆盖率（∼400 ± 55-200 ± 30 Ab/NP）的 AuNP/Ab 层修饰电极。发现抗原偶联的速率与抗体密度以及结合亲和力的变化一致。在抗原浓度较宽的范围内（6.2ng/mL 至 0.12fg/mL），检测下限低至飞摩尔水平（0.25fM/mL）。使用化学计量学方法和响应面法（RSM）进一步优化了影响表位-抗体相互作用的变量。
