Graphene oxide (GO), with its exceptional physical and chemical properties and biocompatibility, holds a tremendous potential for sensing applications. In this study, GO, acting both as the electron-transfer bridge and the signal reporter, was attached on the interface to develop a label-free electrochemical nanosandwich device for detection of interleukin-6 (IL-6). First, a single layer of GO was covalently modified on gold electrodes, followed by attachment of anti-IL-6 capture antibody to form the sensing interface. The 4-aminophenyl phosphorylcholine was further attached to the surface of GO to minimize nonspecific protein adsorption. For reporting the presence of analyte, the anti-IL-6 detection antibody was covalently modified to the GO, which has been integrated with the redox probe Nile blue (NB). Finally, a nanosandwich assay was fabricated on gold surfaces for detection of IL-6 on the basis of the electrochemical signal of NB. The prepared nanosandwiches demonstrated high selectivity and stability for detection of IL-6 over the range of 1–300 pg mL–1 with the lowest detectable concentration of 1 pg mL–1. The device was successfully used for monitoring of IL-6 secretion in RAW cells and live mice. By tailoring the GO surface with functional components, such devices were able to detect the analyte in vivo without causing inflammatory response.Keywords: aryldiazonium salt chemistry; cytokines; graphene oxides; in vivo detection; nanosandwich device;

Aryldiazonium salts as coupling agents for surface chemistry have evidenced their wide applications for the development of sensors. Combined with advances in nanomaterials, current trends in sensor science and a variety of particular advantages of aryldiazonium salt chemistry in sensing have driven the aryldiazonium salt-based sensing strategies to grow at an astonishing pace. This review focuses on the advances in the use of aryldiazonium salts for modifying interfaces in sensors and biosensors during the past decade. It will first summarize the current methods for modification of interfaces with aryldiazonium salts, and then discuss the sensing applications of aryldiazonium salts modified on different transducers (bulky solid electrodes, nanomaterials modified bulky solid electrodes, and nanoparticles). Finally, the challenges and perspectives that aryldiazonium salt chemistry is facing in sensing applications are critically discussed.Keywords: aryldiazonium salts; interface fabrication; nanomaterials; nanosensors; sensing applications;

In this study, a label-free electrochemical immunosensor was developed for detection of cytokine tumor necrosis factor-alpha (TNF-α). First, AuNPs loaded reduced graphene oxides nanocomposites (RGO-ph-AuNP) were prepared, and then, a mixed layer of 4-carbxyphenyl and 4-aminophenyl phosphorylcholine (PPC) was modified to the surface of AuNPs for the subsequent modification of anti-TNF-α capture antibody (Ab1) to form the capture surface (Au-RGO-ph-AuNP-ph-PPC(-ph-COOH)) for the analyte TNF-α with the antifouling property. For reporting the presence of analyte, the anti-TNF-α detection antibody (Ab2) was modified to the graphene oxides which have been modified with the 4-ferrocenylaniline through diazonium chemistry to form Ab2-GO-ph-Fc. Then, a sandwich assay was formed on gold surfaces for the quantitative detection of TNF-α based on the electrochemical signal of ferrocene. X-ray photoelectron spectra (XPS), transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FT-IR), UV–vis, and electrochemistry were used for characterization of the stepwise fabrications on the interface. The prepared electrochemical immunosensor was successfully used for the detection of TNF-α over the range of 0.1–150 pg mL–1. The lowest detection limit of this immunosensor is 0.1 pg mL–1 TNF-α in 50 mM phosphate buffer at pH 7.0. The fabricated immunosensor provided high selectivity and stability and can be used to detect TNF-α secreted by live BV-2 cells with comparable accuracy to enzyme-linked immunosorbent assay (ELISA) but with lower limit of detection.

•Nanocomposites based on GO and AuNPs were prepared and anchored on the electrode surfaces covalently to form a stable sensing interface.•The anti-cTnI detection antibody was immobilized on GO tailored with ferrocene molecules, functioning as the signal reporter for the detection of cTnI.•The detectable concentration of cTnI is 0.05 ng mL-1 in buffer with the assay time of less than 5 min.•The herein simple and novel approach for fabrication of AuNP and graphene based platform is promising for future fabrication of point-of-care devices.A stable label-free amperometric immunosensor is presented based on gold nanoparticles and graphene oxide nanocomposites for detection of cardiac troponin-I in the early diagnosis of myocardial infarction. For designing of the sensing platform, firstly the nanocomposites based on GO and AuNPs were prepared and anchored on electrode surfaces. The formed nanocomposites provided a platform with big surface area for loading anti-cTnI capture antibody, and worked as a bridge for fast electron transfer subsequently increased the sensitivity. Moreover, the linkages between AuNP, GO, and electrodes were based on covalent bonding by aryldiazonium salt coupling chemistry, which favors the stability of the sensing interface. Finally, the anti-cTnI detection antibody was immobilized on GO tailored with ferrocene molecules, functioning as the signal reporter for the detection of cTnI. The modification process was monitored using electrochemistry, SEM, XPS. The herein immunosensor demonstrates a good selectivity and high sensitivity against human-cTnI, and is capable of detecting cTnI at concentrations as low as 0.05 ng mL−1, which is 100 times lower than that possible by conventional methods. It is potential to design the portable sensing platform based on AuNPs and GO nanocomposites for future point-of-care diagnostics.

AuNP decorated GO nanocomposites (GO-Ph-AuNP) via aryldiazonium salt chemistry have been successfully prepared, which can be used as the immobilization matrix for loading glucose oxidase (GOx) towards a sensitive glucose sensor. The fabricated nanocomposite was characterized by field emission scanning electron microscopy, UV-Vis and electrochemistry. The direct electrochemistry of GOx was successfully realized on GO-Ph-AuNP modified GC electrodes with a heterogeneous electron transfer rate constant of 8.3 s−1, revealing fast direct electron transfer of GOx. The GOx immobilized on GO-Ph-AuNP nanocomposite modified electrode retained good electrocatalytic activity toward glucose over a linear concentration range from 0.3 to 20 mM with the sensitivity of 42 μA mM−1 cm−2 and enzyme turnover rate of 112 s−1. In addition, the fabricated biosensor was capable of monitoring glucose consumption by live cells.

This paper reports an electrochemical sensor based on the covalent anchoring of aryldiazonium salt modified AuNPs to gold electrodes for the sensitive detection of cadmium ions (Cd2+). AuNPs modified with 4-nitrophenyl and 4-carboxylphenyl were first prepared, which were then immobilized in situ on gold electrodes to achieve Au–Ph–AuNP modified interfaces. Glutathione (GSH) was finally modified to Au–Ph–AuNP by amide bonding to achieve an Au–Ph–AuNP–GSH sensing interface, which was used to detect Cd2+ over the concentration range from 0.1 nM to 100 nM with a detection limit of 0.1 nM. This electrochemical sensor demonstrated high stability and sensitivity to Cd2+. It has potential use in an in situ anchoring AuNP strategy to fabricate portable devices for the onsite monitoring of trace amounts of heavy metals.

•AuNPs can be covalently anchored to GC surfaces by one-step strategy.•AuNPs were used as both electronic bridges and signal amplifiers in an immunosensor.•The immunosensor can be used for detection of BoNT/A in both buffers and milks.•The detection limit of BoNT/A is 1 pg mL−1 in buffer with the assay time of 10 min.•The herein sensing strategy is promising for designing point-of-care devices.This work introduced an efficient approach for modification of AuNPs with multicomponents by diazonium salt couplings. The multifunctionalized AuNPs with protruding functional groups that allow simple bioconjugation to large amounts of biomolecules have been successfully used as electronic bridges and signal amplifiers for an electrochemical immunosensor towards the detection of BoNT/A. The one-step anchoring AuNPs strategy has greatly increased the efficiency for attachment of biomolecules and subsequently increased the sensitivity. Sensitivity was further amplified by preparation of bioconjugates particles containing horseradish peroxidase (HRP) labels along with detection antibodies (AbL) attached to AuNPs. The immunosensor can be used for the detection of BoNT/A over the range of 4–35 pg mL−1 with the lowest detection limit of 1 pg mL−1 and assay time of 10 min. The herein sensing strategy is rapid, robust, selective, sensitive, and is promising for future fabrication of point-of-care devices.

•A novel multianalyte electrochemical immunosensor based on the assembly of patterned SWNTs on GC substrates is presented.•Forest of SWNTs can be patterned on GC substrates by C3C bonding using micro contact printing (MCP).•The fabricated immunosensor can be used for simultaneous detection of endosulfan and paraoxon with high sensitivity.•The fabricated immunosensor array demonstrates high repeatability, reproducibility, stability and selectivity towards the pesticides detection.A novel multianalyte electrochemical immunosensor based on the assembly of patterned SWNTs on glassy carbon (GC) substrates was developed for simultaneous detection of endosulfan and paraoxon. Based on aryldiazonium salt chemistry, forest of SWNTs can be patterned on GC substrates by C3C bonding using micro contact printing (MCP), which provides an interface showing efficient electron transfer between biomolecules and electrodes. Then redox molecules FDMA and PQQ can be attached to the SWNTs, respectively followed by the attachment of specific epitopes and antibodies. The modified sensing surfaces were characterized by XPS, SEM, AFM and electrochemistry. Based on the current change of specific redox probes, the fabricated immunosensor array can be used for simultaneous detection of endosulfan and paraoxon by a displacement assay. In phosphate buffer solution (50 mM, pH 7.0), there is a linear relationship between electrochemical signal of FDMA and the concentration of endosulfan over the range of 0.05–100 ppb with a detection limit of 0.05 ppb; the linear range between electrochemical signal of PQQ and the concentration of paraoxon is 2–2500 ppb with a detection limit of 2 ppb. The immunosensor array demonstrates high repeatability, reproducibility, stability and selectivity for the detection of endosulfan and paraoxon.

•A sensitive H2O2 biosensor based on aryldiazonium salt and SWNTs is presented.•SWNTs are covalently attached on diazonium salt modified interface by C–C bonding.•Enzyme HRP is covalently attached to SWNTs/PEG modified interface.•The H2O2 biosensor shows high sensitivity, good reproducibility and high stability.•H2O2 can be detected with a detection limit of 10 nM.A stable and sensitive hydrogen peroxide (H2O2) biosensor based on aryldiazonium salt and SWNTs modified gold electrodes is reported. SWNTs were covalently anchored to the mixed monolayer of phenyl and 4-aminophenyl in molar ratio of 1:1 through aryldiazonium salt reaction to form stable C–C bonding. PEG molecules were introduced to the interface to resist non-specific protein adsorption. Covalent attachment of HRP to SWNTs allowed direct electron transfer to the redox protein with a rate constant of 28.6 ± 1.9 s−1, indicating a specific interaction between SWNTs and HRP. The covalently attached SWNTs facilitate the electrical coupling between protein and electrodes. The covalently immobilized HRP retained its catalytic activity by the enzyme responding to the addition of H2O2. The SWNTs/PEG/HRP modified sensing interface can be used for the detection of H2O2 in the range of 0.01–24 μM with a detection limit of 10 nM. Comparing to the sensing system in which HRP was physically adsorbed on the interface without the assembly of PEG, the performance of the SWNTs/PEG/HRP sensing interface has been significantly improved. The so fabricated biosensor exhibited high sensitivity, good reproducibility, and long-term stability, and can be used for the detection of H2O2 in real samples with good recovery.

A label-free immunosensor based on SWNTs modified GC electrodes has been developed for the direct detection of paraoxon. Based on aryldiazonium salt chemistry, forest of SWNTs can be vertically aligned on mixed monolayers of aryldiazonium salt modified GC electrodes by CC bonding, which provides an interface showing efficient electron transfer between biomolecules. PEG molecules were introduced to the interface to resist non-specific protein adsorption. Ferrocenedimethylamine (FDMA) was subsequently attached to the ends of SWNTs through the amide bonding followed by the attachment of epitope i.e., paraoxon hapten to which a paraoxon antibody would bind. This immunosensor shows good selectivity and high specificity to paraoxon, and is functional for the detection of paraoxon in both laboratory and field by a displacement assay. There is a linear relationship between electrochemical signal of FDMA and the concentration of paraoxon over the range of 2–2500 ppb with a lowest detected limit of 2 ppb in 0.1 M phosphate buffer at pH 7.0. The SWNTs based amperometric immunosensor provides an opportunity to develop the sensing system for on-site sensitive detection of a spectrum of insecticides.Highlights► A label-free immunosensor based on SWNTs for detection of paraoxon is presented. ► SWNTs are vertically aligned on diazonium salt modified interface by CC bonding. ► The immunosensor shows high specificity, selectivity, and sensitivity to paraoxon. ► Paraoxon can be detected by a displacement assay with a detection limit of 2 ppb. ► The sensing system has potential for on-site detection of a spectrum of pesticides.

A glassy carbon substrate was covalently modified with a mixed layer of 4-aminophenyl and phenyl via in situ electrografting of their aryldiazonium salts in acidic solutions. Single-walled carbon nanotubes (SWNTs) were covalently and vertically anchored on the electrode surface via the formation of amide bonds from the reaction between the amines located on the modified substrate and the carboxylic groups at the ends of the nanotubes. Ferrocenedimethylamine (FDMA) was subsequently attached to the ends of SWNTs through amide bonding followed by the attachment of an epitope, i.e., endosulfan hapten to which an antibody would bind. Association or dissociation of the antibody with the sensing interface causes a modulation of the ferrocene electrochemistry. Antibody-complexed electrodes were exposed to samples containing spiked endosulfan (unbound target analyte) in environment water and interrogated using the square wave voltammetry (SWV) technique. The modified sensing surfaces were characterized by atomic force microscopy, XPS, and electrochemistry. The fabricated electrochemical immunosensor can be successfully used for the detection of endosulfan over the range of 0.01–20 ppb by a displacement assay. The lowest detection limit of this immunosensor is 0.01 ppb endosulfan in 50 mM phosphate buffer at pH 7.0.

This paper reports an electrochemical sensor based on the covalent anchoring of aryldiazonium salt modified AuNPs to gold electrodes for the sensitive detection of cadmium ions (Cd2+). AuNPs modified with 4-nitrophenyl and 4-carboxylphenyl were first prepared, which were then immobilized in situ on gold electrodes to achieve Au–Ph–AuNP modified interfaces. Glutathione (GSH) was finally modified to Au–Ph–AuNP by amide bonding to achieve an Au–Ph–AuNP–GSH sensing interface, which was used to detect Cd2+ over the concentration range from 0.1 nM to 100 nM with a detection limit of 0.1 nM. This electrochemical sensor demonstrated high stability and sensitivity to Cd2+. It has potential use in an in situ anchoring AuNP strategy to fabricate portable devices for the onsite monitoring of trace amounts of heavy metals.