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.

Non-Langmuir Kinetics of DNA Surface Hybridization / L. Vanjur, T. Carzaniga, L. Casiraghi, M. Chiari, G. Zanchetta, M. Buscaglia. - In: BIOPHYSICAL JOURNAL. - ISSN 0006-3495. - 119:5(2020 Sep 01), pp. 989-1001. [10.1016/j.bpj.2020.07.016]

Non-Langmuir Kinetics of DNA Surface Hybridization

L. Vanjur
Primo
Formal Analysis
;
T. Carzaniga
Secondo
Investigation
;
G. Zanchetta
Penultimo
Writing – Review & Editing
;
M. Buscaglia
Ultimo
Supervision
2020-09-01

Abstract

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.
Settore FIS/07 - Fisica Applicata(Beni Culturali, Ambientali, Biol.e Medicin)
Piattaforma per l’identificazione di target di rilevanza farmacologica per il trattamento di patologie del sistema nervoso e oncologiche ad elevato bisogno di cura (NeOn)
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2434/763270
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