Revealing the Critical Role of Probe Grafting Density in Nanometric Confinement in Ionic Signal via an Experimental and Theoretical Study

Abstract

The grafting density of probes at sensor interface plays a critical role in the performance of biochemical sensors. However, compared with macroscopic interface, the effects of probe grafting density at nanometric confinement are rarely studied due to the limitation of precise grafting density regulation and characterization at the nanoscale. Here, we investigate the effect from the grafting density of DNA probes on ionic signal for nucleic acid detection in a cylindrical nanochannel array (with diameter of 25 nm) by combing experiments and theories. We set up a theoretical model of charge distribution from close to inner wall of nanochannels at low probe grafting density to spreading in whole space at high probe grafting density. The theoretical results fit well with the experimental results. A reverse of ionic output from signal-off to signal-on occurs with increasing probe grafting density. Low probe grafting density offers a high current change ratio that is further enhanced using long-chain DNA probes or the electrolyte with a low salt concentration. This work develops an approach to enhance performance of nanochannel-based sensors and explore physicochemical properties in nanometric confines.

Revealing Ionic Signal Enhancement with Probe Grafting Density on the Outer Surface of Nanochannels

“…Taking the current at + 2 V for comparison, the change rate of ion current (β = ( I probe+target – I probe )/ I probe ) was calculated. The β value reflects the conversion efficiency from target characteristics to the ionic signal, related to the sensitivity and specificity of nanopore/nanochannel analyses. ,,,− As shown in Figure c, the β increased with the grafting density of PNA, which was further confirmed by four parallel experiments using different concentrations of DNA targets.…”

“…The Poisson–Nernst–Planck (PNP) equations were used to numerically simulate the ionic current of the nanochannels with different grafting densities of PNA probes on the outer surface, in which the measured values of the grafting density and distribution of PNA were used. Commonly, the changing of the pore size reflects the change of the steric hindrance in numerical simulations. ,− However, it cannot reflect the change of the probe grafting density (Figure S3). With the construction of the geometric model for the independent probe (Figure a), the physical meaning of each probe has been taken into account, which precisely reflects the change of the probe grafting density.…”

“…This effect was only roughly investigated through fluorescence characterization and numerical simulations in our previous work. 41 Now, high-resolution characterization methods, including surface charge and steric hindrance, are available for the probes on the outer surface of nanochannels. 35 In this study, the electroneutral peptide nucleic acids (PNAs) acting as probes for DNA targets were immobilized on the outer surface of Au-deposited poly(ethylene terephthalate) (PET) nanochannels.…”

“…As a significant parameter of the sensing interface, the influence of the probe grafting density in nanopores or nanochannels has been ignored for a long time because it is difficult to precisely control and characterize the distribution of probes inside the nanoconfined cavities. This effect was only roughly investigated through fluorescence characterization and numerical simulations in our previous work . Now, high-resolution characterization methods, including surface charge and steric hindrance, are available for the probes on the outer surface of nanochannels …”

Exponential Increase in an Ionic Signal: A Dominant Role of the Space Charge Effect on the Outer Surface of Nanochannels

“…46 Thus, it could potentially tackle the inefficiencies of the ionic signal conversion resulted from the offset between the space charge and steric effect of targets. 47 However, the space charge and steric effect of the charged targets on the outer surface on the ionic signal have not been separately and quantitatively demonstrated. The corresponding signaling mechanisms are not explicit.…”