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Non-Langmuir kinetics of DNA surface hybridization
Biophysical Journal ( IF 3.2 ) Pub Date : 2020-09-01 , DOI: 10.1016/j.bpj.2020.07.016
Luka Vanjur 1 , Thomas Carzaniga 1 , Luca Casiraghi 1 , Marcella Chiari 2 , Giuliano Zanchetta 1 , Marco Buscaglia 1
Affiliation  

Hybridization of complementary single strands of DNA represents a very effective natural molecular recognition process widely exploited for diagnostic, biotechnology, and nanotechnology applications. A common approach relies on the immobilization on a surface of single-stranded DNA probes that bind complementary targets in solution. However, despite the deep knowledge on DNA interactions in bulk solution, the modeling of the same interactions on a surface are still challenging and perceived as strongly system dependent. Here, we show that a two-dimensional analysis of the kinetics of hybridization, performed at different target concentrations and probe surface densities by a label-free optical biosensor, reveals peculiar features inconsistent with an ideal Langmuir-like behavior. We propose a simple non-Langmuir kinetic model accounting for an enhanced electrostatic repulsion originating from the surface immobilization of nucleic acids and for steric hindrance close to full hybridization of the surface probes. The analysis of the kinetic data by the model enables quantifying the repulsive potential at the surface, as well as retrieving the kinetic parameters of isolated probes. We show that the strength and the kinetics of hybridization at large probe density can be improved by a three-dimensional immobilization strategy of probe strands with a double-stranded linker.

中文翻译:


DNA 表面杂交的非 Langmuir 动力学



DNA 互补单链的杂交代表了一种非常有效的天然分子识别过程,广泛用于诊断、生物技术和纳米技术应用。一种常见的方法依赖于单链 DNA 探针表面的固定化,这些探针与溶液中的互补靶标结合。然而,尽管对本体溶液中 DNA 相互作用有了深入的了解,但表面上相同相互作用的建模仍然具有挑战性,并且被认为强烈依赖于系统。在这里,我们表明,通过无标记光学生物传感器在不同的目标浓度和探针表面密度下进行的杂交动力学的二维分析揭示了与理想的朗缪尔样行为不一致的独特特征。我们提出了一个简单的非朗缪尔动力学模型,解释了源自核酸表面固定的增强静电排斥以及接近表面探针完全杂交的空间位阻。通过模型对动力学数据的分析可以量化表面的排斥势,并检索孤立探针的动力学参数。我们表明,通过具有双链接头的探针链的三维固定策略可以提高大探针密度下杂交的强度和动力学。
更新日期:2020-09-01
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