The Fabrication of a Probe-Integrated Electrochemiluminescence Aptasensor Based on Double-Layered Nanochannel Array with Opposite Charges for the Sensitive Determination of C-Reactive Protein

Abstract

The effective and sensitive detection of the important biomarker, C-reactive protein (CRP), is of great significance in clinical diagnosis. The development of a convenient and highly sensitive electrochemiluminescence (ECL) aptasensor with an immobilized emitter probe is highly desirable. In this work, a probe-integrated ECL aptamer sensor was constructed based on a bipolar silica nanochannel film (bp-SNF) modified electrode for the highly sensitive ECL detection of CRP. The bp-SNF, modified on an ITO electrode, consisted of a dual-layered SNF film, including the negatively charged inner SNF (n-SNF) and the outer SNF with a positive charge and amino groups (p-SNF). The ECL emitter, tris(bipyridine) ruthenium (II) (Ru(bpy)32+), was stably immobilized in a nanochannel of bp-SNF using the dual electrostatic interactions with n-SNF attracting and p-SNF repelling. The amino groups on the outer surface of bp-SNF were aldehyde derivatized, allowing for the covalent immobilization of recognitive aptamers (5′-NH2-CGAAGGGGATTCGAGGGGTGATTGCGTGCTCCATTTGGTG-3′), leading to the recognition interface. When CRP bound to the aptamer on the recognition interface, the formed complex increased the interface resistance and reduced the diffusion of the co-reactant tripropylamine (TPA) into the nanochannels, leading to a decrease in the ECL signal. Based on this mechanism, the constructed aptamer sensor could achieve a sensitive ECL detection of CRP ranging from 0.01 to 1000 ng/mL, with a detection limit (DL) of 8.5 pg/mL. The method for constructing this probe-integrated ECL aptamer sensor is simple, and it offers a high probe stability, good selectivity, and high sensitivity.

Solid Electrochemiluminescence Sensor by Immobilization of Emitter Ruthenium(II)tris(bipyridine) in Bipolar Silica Nanochannel Film for Sensitive Detection of Oxalate in Serum and Urine

“…On the one hand, the immobilization of ECL-emitting molecules prevents their dissolution, drift, or loss during operation, ensuring the stability of the luminescent material and thereby enhancing the reliability of the sensor. Moreover, solid-state ECL systems contribute to increased signal intensity [ 29 ]. On the other hand, the reaction between the ECL emitter and the electrode is facilitated by tightly fixing the ECL emitter to the electrode surface, improving sensitivity.…”

“…Utilizing porous molecular sieve film to modify electrodes provides an effective method for constructing solid-state ECL systems. Among them, silica nanochannel array film (SNA) has attracted considerable attention [ 29 , 30 ]. Specifically, SNA is prepared through a sol–gel process using surfactant micelles (SM) as templates, and it exhibits not only high mechanical and thermal stability and biocompatibility but also advantages such as uniform distribution of nanochannels (typically, 2–3 nm), tunable pore sizes (enlarged by introducing pore-expanders), and a high specific surface area [ 38 , 39 , 40 , 41 ].…”

Vertical Silica Nanochannels and o-Phenanthroline Chelator for the Detection of Trace Fe(II)

“…Molecules with different characteristics can show distinct mass transport processes and physical phenomena at the nanoscale, which has been employed for the construction of novel electrochemical and electrochemiluminescence sensors with high sensitivity. , For example, when the size of charged nanochannels or nanopores is comparable to the Debye length (a few or tens of nanometers) in solution, counterions will be electrostatically attracted due to their interaction with the electrical double layer (EDL) near the charged surface of nanochannels or nanopores. − In addition, molecules confined in ultrasmall volume containers can be more easily detected compared to those in bulk solution, which arises from the ultrasmall volume and greatly increased concentration of molecules. − Vertically ordered mesoporous silica film (VMSF) consisting of uniform (commonly 2–3 nm in diameters) and numerous (10 12 pores cm –2 ) nanopores and ultrathin thickness (∼100 nm as usual) has been utilized to modify the electrode for sensitive and antifouling detection of various analytes of interests in real samples, such as metal ions, biomarkers, cells, and drug molecules in complex samples. − Thanks to the deprotonation of abundant silanol groups (p K a ∼ 2) inside nanochannels, ultrasmall volume (∼0.48 zL) of a nanochannel, and adjustable hydrophobic surface, VMSF has become an effective permselective electrode material in terms of charge, size, and complexation effects, which has aroused intensive fundamental research and sensing work so far. − To the best of our knowledge, exploitation of chelating agents for promoting the confined effect inside nanochannels of VMSF and further increasing the analytical performance has not yet been reported.…”

Highly Sensitive Electrochemical Determination of Butylated Hydroxyanisole in Food Samples Using Electrochemical-Pretreated Three-Dimensional Graphene Electrode Modified with Silica Nanochannel Film

“…VMSF is composed of a vertical array of nanochannels perpendicular to the electrode, uniform nanochannel sizes (typically 2-3 nm in diameter), high pore density (up to 10 12 /cm 2 ), adjustable thickness (usually 50-200 nm), and good chemical stability [43][44][45]. Compared to the morphologies of other porous silica materials, these characteristics provide VMSF with rapid mass transfer capabilities and size and electrostatic sieving capabilities at the molecular level [46][47][48][49][50]. For instance, it has been proven that the monomer of microperoxidase-11 (MP-11, Mw 1861, size: 1.1 × 1.7 × 3.3 nm), a hemecontaining peptide with only 11 amine acids, can enter the nanochannels of VMSF through adsorption.…”