Immunosensing Method Research Articles

Perylene tetracarboxylic acid (PTA)-based type II heterojunction sensing platform for signal-on photoelectrochemical detection of CEA

The construction of robust photoelectrochemical (PEC) transducing platforms mediates the feasibility and performance of the PEC biosensors. In this work, perylene tetracarboxylic acid (PTA) nanoassemblies were exploited as robust semiconductors to form type II heterojunctions with In2O3. The prepared organic/inorganic heterojunctions show high PEC transducing performances. The carboxyl groups of PTA avoid the additional surface functionalization for covalent immobilization of capturing elements which will diminish the transducing performance. This enables the facile construction of PEC immunosensing platforms. More importantly, Ag@CeO2 nanoparticles were used as energy level matching nanolabels to develop a signal-on PEC immunosensing method. A linear response from 0.1 pg/mL to 100 ng/mL with a detection limit of 63.1 fg/mL was achieved for a typical protein biomarker carcinoembryonic antigen (CEA).

A novel electrochemical immunosensor based on SnS2/NiCo metal-organic frameworks loaded with gold nanoparticles for cortisol detection

In this work, a label-free electrochemical immunosensor using tin sulfide/nickel cobalt metal-organic frameworks (SnS2/NiCo MOFs) was established for the sensitive detection of cortisol. First, SnS2/NiCo MOFs were synthesized by doping SnS2 with NiCo MOF nanocubes by a hydrothermal method. Then, gold nanoparticles (AuNPs) were grown in situ on SnS2/NiCo MOFs for electrochemical detection. The use of SnS2/NiCo MOFs promoted the electron transfer rate of AuNPs and enhanced the electrochemical sensing performance of AuNPs@SnS2/NiCo MOFs-modified electrodes. The large specific surface area of AuNPs@SnS2/NiCo MOFs provides more active sites for antibody loading. After the prepared immunosensor was incubated with the target analyte, cortisol, the electron transfer impedance increased and the amperometric response decreased, thus establishing a highly sensitive immunosensing method. The sensor had a linear range of 100 fg/mL to 100 ng/mL and a low detection limit of 29 fg/mL. The sensor showed good accuracy and practicability and could be used for the determination of cortisol in saliva.

Trace Determination of Grouper Nervous Necrosis Virus in Contaminated Larvae and Pond Water Samples Using Label-Free Fiber Optic Nanoplasmonic Biosensor.

We developed a fast (<20 min), label-free fiber optic particle plasmon resonance (FOPPR) immunosensing method to detect nervous necrosis virus (NNV), which often infects high-value economic aquatic species, such as grouper. Using spiked NNV particles in a phosphate buffer as samples, the standard calibration curve obtained was linear (R2 = 0.99) and the limit of detection (LOD) achieved was 2.75 × 104 TCID50/mL, which is superior to that obtained using enzyme-linked immunosorbent assay (ELISA). By using an enhancement method called fiber optic nanogold-linked immunosorbent assay (FONLISA), the LOD can be further improved to <1 TCID50/mL, which is comparable to that found by the conventional qPCR method. Employing the larvae homogenate samples of NNV-infected grouper, the results obtained by the FOPPR biosensor agree with those obtained by the quantitative polymerase chain reaction (qPCR) method. We also examined pond water samples from an infected container in an indoor aquaculture facility. The lowest detectable level of NNV coat protein was found to be 0.17 μg/mL, which is one order lower than the LOD reported by ELISA. Therefore, we demonstrated the potential of the FOPPR biosensor as an outbreak surveillance tool, which is able to give warning indication even when the trend of larvae death toll increment is still not clear.

Photocurrent quenching by competitive consumption of surface electron donor and light absorption for immunosensing

This work designs a competitive consumption strategy of surface electron donor and light absorption for quenching the photocurrent of ZnSnO3 nanocubes/BiOI nanoarrays/polydopamine (ZnSnO3 NCs/BiOI NAs/PDA) as a photoactive material. This material can be formed on electrode surface by successive coating and deposition to provide a substrate for immobilization of capture antibody, and producing strong photocurrent in the presence of ascorbate acid as a surface electron donor due to the well matching structure of band gaps between ZnSnO3 NCs and BiOI NAs, the excellent light absorption ability and high photo-electron conversion efficiency of BiOI NAs and PDA, and the accelerated electron transfer. Using ascorbate oxidase loaded dopamine-melanin nanosphere (DAM-AAO) as a label of the signal (secondary) antibody, the sandwich-type immunoreaction leads to dual photocurrent quenching of the label through the competitive consumption of ascorbate acid with enzymatic oxidation and the light absorption by DAM nanosphere. Thus, a sensitive “On-Off” photoelectrochemical (PEC) immunosensing method is constructed for the analysis of neuron specific enolase (NSE). The proposed method shows a detection range of 0.1 pg/mL −50 ng/mL and a detection limit of 0.03 pg/mL. The excellent performance and the recovery text demonstrated the practicability of the designed strategy and label in immunoassay of different protein biomarkers.

Peroxidase-Mimicking Activity of Biogenic Gold Nanoparticles Produced from Prunus nepalensis Fruit Extract: Characterizations and Application for the Detection of Mycobacterium bovis.

In the present study, a facile, eco-friendly, and controlled synthesis of gold nanoparticles (Au NPs) using Prunus nepalensis fruit extract is reported. The biogenically synthesized Au NPs possess ultra-active intrinsic peroxidase-like activity for the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB) in the presence of H2O2. Chemical analysis of the fruit extract demonstrated the presence of various bioactive molecules such as amino acids (l-alanine and aspartic acids), organic acids (benzoic acid and citric acid), sugars (arabinose and glucose), phenolic acid, and bioflavonoids (niacin and myo-inositol), which likely attributed to the formation of stable biogenic Au NPs with excellent peroxidase-mimicking activity. In comparison with the natural horseradish peroxidase (HRP) enzyme, the biogenic Au NPs displayed a 9.64 times higher activity with regard to the reaction velocity at 6% (v/v) H2O2, presenting a higher affinity toward the TMB substrate. The Michaelis–Menten constant (KM) values for the biogenic Au NPs and HRP were found to be 6.9 × 10–2 and 7.9 × 10–2 mM, respectively, at the same concentration of 100 pM. To investigate its applicability for biosensing, a monoclonal antibody specific for Mycobacterium bovis (QUBMA-Bov) was directly conjugated to the surface of the biogenic Au NPs. The obtained results indicate that the biogenic Au NPs-QUBMA-Bov conjugates are capable of detecting M. bovis based on a colorimetric immunosensing method within a lower range of 100 to 102 cfu mL–1 with limits of detection of ∼53 and ∼71 cfu mL–1 in an artificial buffer solution and in a soft cheese spiked sample, respectively. This strategy demonstrates decent specificity in comparison with those of other bacterial and mycobacterial species. Considering these findings together, this study indicates the potential for the development of a cost-effective biosensing platform with high sensitivity and specificity for the detection of M. bovis using antibody-conjugated Au nanozymes.

Quenching of tungsten-based polyoxometalate nanoclusters on electrochemiluminescence emission of luminol loaded CeVO4/Au for immunoassay of protein

A quenching-typed electrochemiluminescence (ECL) immunosensing method was proposed for protein detection by the redox reaction of tungsten-based polyoxometalate nanoclusters (W-POM NCs) with free radical (O2•–) as the intermediate of co-reactant H2O2. The immunosensor was constructed by co-immobilizing luminol loaded CeVO4/Au nanohybrids and capture antibody on an indium tin oxide electrode, which showed strong ECL emission in the presence of H2O2 due to the catalysis of CeVO4/Au toward its electro-oxidation to produce O2•– via the reversible conversion between Ce3+ and Ce4+. The W-POM NCs were encapsulated in the uniformly sized amino-silica nanosphere (SiO2) to act as a label (W-POM@SiO2) of the secondary antibody. Upon the sandwich-typed immunoreactions with the target, the W-POM NCs were introduced onto the immunosensor surface to reduce O2•–, and thus quenched the ECL emission of luminol-H2O2 system. Using neuron-specific enolase (NSE) as a target, the proposed method showed a detection limit of 0.19 pg/mL and detectable range of 0.5 pg/mL to 18 ng/mL. The excellent performance of the proposed “ON-OFF” strategy demonstrated that the W-based quenching offers an effective tag for immunoassay of protein biomarkers.

Enzyme-catalyzed deposition of polydopamine for amplifying the signal inhibition to a novel Prussian blue-nanocomposite and ultrasensitive electrochemical immunosensing

The uncontrollable synthesis of Prussian blue (PB) and its weak stability toward OH– are great challenges affecting its electrochemical biosensing application. Herein we utilize the unique properties of chitosan (CS) to realize the facile and controllable synthesis of a CS-PB nanocomposite and combine it with the urease-catalyzed deposition of polydopamine (PDA) for amplifying the electrochemical signal inhibition of PB to develop a novel immunosensing method for protein detection. The immunosensor was constructed on a CS-PB modified electrode, and a urease-functionalized silica nanoprobe was prepared for tracing its sandwich immunoassay toward the model analyte of carcinoembryonic antigen. Besides the electrochemical impedance effect of the quantitatively captured nanoprobes, their enzymatic reaction can release numerous OH– to destroy the PB crystals and also induce the PDA deposition onto the immunosensor. These caused drastic electrochemical signal inhibition to PB. Based on the above multi-signal amplification mechanism, the method exhibits a very low detection limit of 0.042 pg mL–1 along with a very wide linear range of six-order of magnitude. In addition, the CS-PB based immunosensor has excellent specificity, repeatability, stability and reliability. Thus this PB nanocomposite and the proposed electrochemical immunosensing method reveal a promising potential for future applications.

Cysteine Modification of Glutathione-Stabilized Au Nanoclusters to Red-Shift and Enhance the Electrochemiluminescence for Sensitive Bioanalysis.

Screening new electrochemiluminescence (ECL) emitters for the design of sensitive detection strategies with even long emission wavelength is intensively anticipated in ECL evolution. Herein, a promising modification strategy for improving the ECL performance of Au nanoclusters (AuNCs) as a water-soluble luminophore was proposed. Upon the introduction of l-cysteine (l-Cys) onto the surface of glutathione (GSH)-stabilized AuNCs (GSH-AuNCs), the dual-thiol bond between l-Cys and GSH was formed to limit the intramolecular motion and nonradiative relaxation of the excited state from the capping agents, which resulted in the enhancement of monochromatic ECL emission of GSH-AuNCs with a red-shifted wavelength. By utilizing triethylamine as a coreactant, the ECL of l-Cys/GSH-AuNCs was about 1.5-fold stronger than that of GSH-AuNCs, and the emission wavelength red-shifted from 660 to 780 nm at a relatively low potential, which could decrease the interference in bioassay and the photochemical damage in nondestructive detection. As a proof of application, a sandwich-type immunosensing method for CYFRA 21-1 was proposed with l-Cys/GSH-AuNCs as the signal tag, which displayed a wide linear ranging from 0.2 fg/mL to 2 ng/mL and a limit of detection down to 0.067 fg/mL at 3S/N. This work provides a wonderful strategy for promoting the performance of ECL emitters.

Magnet-assisted electrochemical immunosensor based on surface-clean Pd-Au nanosheets for sensitive detection of SARS-CoV-2 spike protein

Tracking and monitoring of low concentrations of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can effectively control asymptomatic transmission of current coronavirus disease 2019 (COVID-19) in the early stages of infection. Here, we highlight an electrochemical immunosensor for sensitive detection of SARS-CoV-2 antigen marker spike protein. The surface-clean Pd-Au nanosheets as a substrate for efficient sensing and signal output have been synthesized. The morphology, chemical states and excellent stable electrochemical properties of this surface-clean heterostructures have been studied. Functionalized superparamagnetic nanoparticles (MNPs) were introduced as sample separators and signal amplifiers. This biosensor was tested in phosphate buffered saline (PBS) and nasopharyngeal samples. The results showed that the sensor has a wide linear dynamic range (0.01 ng mL−1 to 1000 ng mL−1) with a low detection limit (0.0072 ng mL−1), which achieved stable and sensitive detection of the spike protein. Therefore, this immunosensing method provides a promising electrochemical measurement tool, which can furnish ideas for early screening and the reasonable optimization of detection methods of SARS-CoV-2.

Copper doped terbium metal organic framework as emitter for sensitive electrochemiluminescence detection of CYFRA 21-1

Lanthanide metal organic frameworks (L-MOFs) are emerging as promising electrochemiluminescence (ECL) emitters for bioanalysis. This work proposed a copper doped terbium MOF as a luminescent tag for construction of a “signal-on” ECL immunosensing method. The Tb–Cu-PA MOF was prepared using Tb3+ and Cu2+ ions as metal linkers and m-phthalic acid as bridge ligand, and exhibited strong ECL emission with K2S2O8 as a coreactant. The immunosensor was prepared by immobilizing capture antibody on Pd nanoparticles modified Ni–Co layered double hydroxide (Pd-ZIF-67@LDH) nanoboxes, which showed strong electrocatalytic activity toward the reduction of S2O82− for amplifying the ECL signal. Upon the sandwich-typed immunoreactions, Tb–Cu-PA MOF labeled antibody was introduced onto the immunosensor for sensitive ECL detection of target protein. Using cytokeratin 19 fragment 21–1 (CYFRA21-1), a representative lung cancer biomarker, as target model, the ECL immunosensing method showed a linear range of 0.01–100 ng/mL and a detection limit of 2.6 pg/mL (S/N = 3). This immunosensing strategy highlighted the advances of using luminescent and electroactive MOFs in the developments of highly efficient immunosensors for bioanalysis.

A DNA dendrimer amplified electrochemical immunosensing method for highly sensitive detection of prostate specific antigen

This work designed a DNA dendrimer for the loading of signal molecule and the construction of amplified electrochemical immunosensing method. The DNA dendrimer was self-assembled by the hybridization of one couple of complementary oligonucleotides (DNA and cDNA) that were covalently conjugated to three arms of a Y-shaped cross-linker, tris(2-maleimidoethyl)amine (TMEA) respectively. The immunosensor was prepared by coating chitosan on glassy carbon electrode to covalently immobilize the capture antibody with glutaraldehyde as a linker. After the target protein was captured on the immunosensor, cDNA-labeled secondary antibody was bound on the surface via a sandwiched immunoreaction to introduce the DNA dendrimer onto immunosensor for loading abundant methylene blue as signal molecule, which amplified greatly the amperometric signal for immunoassay. Using prostate specific antigen (PSA) as a model analyte, this proposed method showed a wide linear range from 1 pg mL−1 to 10 ng mL−1 along with a limit of detection down to 0.26 pg mL−1. The designed strategy avoided complex synthesis of signal tags, and possessed excellent performance for analysis of practical samples, thus providing a new avenue for the development of signal amplification strategy and immunoassay methods.

CRISPR/Cas12a-powered immunosensor suitable for ultra-sensitive whole Cryptosporidium oocyst detection from water samples using a plate reader

Waterborne pathogens, such as Cryptosporidium parvum, pose a major threat to public health globally, and this requires screening of drinking and environmental water for low number of contaminating microbes. However, current detection approaches generally require trained experts with sophisticated instruments, and are not suitable for large-scale screening and rapid outbreak response. Recent advances in ultrasensitive CRISPR/Cas-based biosensing continue to expand the range of detectable molecular targets, however single microbes could not be directly detected so far, especially in environmental samples. Here, we report an ultrasensitive CRISPR/Cas12a-powered immunosensing method suitable for microbial detection which links antibody-based recognition with CRISPR/Cas12a-based fluorescent signal amplification through an antibody-DNA conjugate. This approach is shown here to detect whole 4 µm size Cryptosporidium parvum oocysts with a linear range from 6.25 – 1600 oocysts/mL, at a maximum sensitivity of single oocyst per sample. Its potential to apply to various complex sample matrices has also been demonstrated. After sample dilution by factor of 10, we were able to detect 10 oocysts from a back-wash mud samples from water treatment plate. This method uses the same experimental setup (plate reader) as a conventional ELISA assay thus reducing the need for microscopy-based identification of Cryptosporidium, which represents the gold-standard but requires high level expertise and time-consuming manual counting. This work highlights the potential of CRISPR/Cas-based biosensing for water quality assessment and ultrasensitive whole pathogen detection.

An enzyme-free electrochemical immunosensor based on quaternary metallic/nonmetallic PdPtBP alloy mesoporous nanoparticles/MXene and conductive CuCl2 nanowires for ultrasensitive assay of kidney injury molecule-1

Herein, a novel enzyme-free electrochemical immunosensor based on quaternary metallic/nonmetallic PdPtBP alloy mesoporous nanoparticles (MNPs)/MXenes and CuCl2 nanowires (NWs) is reported for detection of kidney injury molecule-1 (KIM-1). PdPtBP MNPs/MXene nanocomposites, the novel signal amplification labels, were successfully synthesized using MXenes with the large surface area and the quaternary PdPtBP MNPs nanozymes with outstanding peroxidase-like catalytic activity. Meanwhile, the combination of high conductive CuCl2 NWs with superior biocompatible gold nanoparticles (AuNPs) is introduced as the substrate to modify the glassy carbon electrode (GCE). Benefiting from the excellent electrochemical performances of PdPtBP MNPs/MXene and CuCl2 NWs, the sandwich-type electrochemical immunosensor showed outstanding analytical performance in the presence of hydrogen peroxide (H2O2). Under the optimized conditions, the electrochemical immunosensor exhibited a good linear response from 0.5 ng/mL to 100 ng/mL with a detection limit of 86 pg/mL. Additionally, the established immunosensing method was successfully applied for the specific detection of KIM-1 in the urine. Thus, the enzyme-free electrochemical immunosensor provides a promising prospect for the detection of KIM-1 in the field of clinical diagnostics.

Nanobody-based immunosensing methods for safeguarding public health.

Immunosensing methods are biosensing techniques based on specific recognition of an antigen–antibody immunocomplex, which have become commonly used in safeguarding public health. Taking advantage of antibody-related biotechnological advances, the utilization of an antigen-binding fragment of a heavy-chain-only antibody termed as 'nanobody' holds significant biomedical potential. Compared with the conventional full-length antibody, a single-domain nanobody retaining cognate antigen specificity possesses remarkable physicochemical stability and structural adaptability, which enables a flexible and efficient molecular design of the immunosensing strategy. This minireview aims to summarize the recent progress in immunosensing methods using nanobody targeting tumor markers, environmental pollutants, and foodborne microbes.

Polyacrylic acid/polyethylene glycol hybrid antifouling interface for photoelectrochemical immunosensing of MDA-MB-231 cells using BiOBr/FeTPPCl/BiOI co-sensitized composite as matrix

An ultrasensitive photoelectrochemical (PEC) antifouling immunosensor was proposed using BiOBr/FeTPPCl/BiOI as matrix and polyacrylic acid (PAA)/polyethylene glycol (PEG) as hybrid antifouling interface for MDA-MB-231 detection. An indium tin oxide (ITO) electrode was successively modified with bismuth oxybromide (BiOBr) with layered crystal structure, which was sensitized with tetraphenyl iron(Ⅲ) porphyrin chloride (FeTPPCl) to widen the absorption range of visible light, and decorated with bismuth oxyiodide (BiOI) nanosheets to improve the PEC response. The joint action of PAA and PEG could offset the adsorption of non-specific proteins on the immunosensing interface for obtaining good antifouling performance. Based on the specific interaction of sialic acid (SA) on the cytomembrane with 3-aminophenylboronic acid (APBA), and the synergistic antifouling effect of PEG and PAA, a simple PEC immunosensing method was developed for the quantitative determination of MDA-MB-231 cells. This proposed antifouling PEC immunosensor exhibited a wide linear range (1 × 102 ∼ 1 × 106 cells·mL-1) and a low detection limit (30 cells·mL-1). The development of antibody-free immunosensors may improve the application for the diagnosis of human breast cancer.

A novel bioassay for the monitoring of carcinoembryonic antigen in human biofluid using polymeric interface and immunosensing method.

Carcinoembryonic antigen (CEA) is a member of a family of cell surface glycoproteins. Recognition of CEA is needed to monitor the physiological status of the patient for treatment and also it is important to assess the severity of the disease. In this work, we reported a novel sandwich-type electrochemical immunosensor based on gold nanoparticles functionalized cysteamine-glutaraldehyde (AuNPs-CysA-GA) and it successfully designed to detection of the CEA biomarker in a human plasma sample. The AuNPs-CysA-GA provides a large surface area for the effective immobilization of CEA antibody, as well as it ascertains the bioactivity and stability of immobilized CEA antigens. Biotinylated-anti-CEA antibody (Ab1) was immobilized on the surface of glassy carbon electrode (GCE) modified AuNPs-CysA-GA. Also, secondary antibody (HRP-Ab2) was costed immobilized to complete the sandwich part of immunosensor. Field emission scanning electron microscope (FE-SEM and EDS), was employed to monitor the sensor fabrication procedure. The immunosensor was used for the detection of CEA using differential pulse voltammetry (DPVs) technique. The proposed interface led to enhancement of accessible surface area for immobilizing high amount of anti-CEA antibody, increasing electrical conductivity, boosting stability, and biocompatibility. Finally, the low limit of quantitation (LLOQ) of the proposed immunosensor was obtained as 7 ng/mL with the linear range of 0.001-5 μg/L. The proposed immunoassay was successfully applied for the monitoring of the CEA in unprocessed human plasma samples. Obtained results paved that the proposed bioassay can be used as a novel bioassay for the clinical diagnosis of cancer based on CEA monitoring.

Label-free electrochemical immunosensor with palladium nanoparticles functionalized MoS2/NiCo heterostructures for sensitive procalcitonin detection

An ultrasensitive label-free electrochemical immunosensor using MoS2/NiCo heterostructures as an efficient signal enhancer was proposed for sensitive detection of procalcitonin. The MoS2/NiCo heterostructures were synthesized by thermally depositing MoS2 on NiCo MOFs nanocubes, which were then functionalized with in situ grown palladium nanoparticles (Pd NPs) for electrochemically catalyze the reduction of hydrogen peroxide. The presence of MoS2/NiCo heterostructures greatly enhanced the electrocatalytic activity of Pd NPs, thus the Pd NPs@MoS2/NiCo modified electrode showed amplified response to hydrogen peroxide. The large specific surface of Pd NPs@MoS2/NiCo greatly improved the loading of antibody for preparation of immunosensor. Upon the incubation of the immunosensor with target procalcitonin, the electron transfer impendence increased, and thus the chronoamperometric response decreased, leading to a highly sensitive immunosensing method for detection of procalcitonin. The proposed method showed a linear range of 0.001–50 ng/mL with a detection limit of 0.36 pg/mL (S/N = 3). It possessed excellent accuracy and good practicality for diagnostics of PCT related diseases, and could be extended for development of other immunosensors for biomarkers detection.

Electrochemical immunosensor for the determination of the cytokine interferon gamma (IFN-γ) in saliva

A simple, fast and sensitive amperometric immunosensing method for the determination of the clinically relevant cytokine interferon gamma (IFN-γ) in saliva complying the requirements demanded for this kind of sample is reported. The target analyte was sandwiched between a specific capture antibody covalently immobilized on a screen-printed electrode functionalized by the diazonium salt grafting of p-aminobenzoic acid, and a biotinylated detector antibody labeled with a streptavidin-horseradish peroxidase conjugate. The amperometric responses measured at - 0.20 V vs Ag pseudo-reference electrode upon addition of hydrogen peroxide in the presence of hydroquinone as the redox mediator allowed a calibration plot with a linear range between 2.5 and 2000 pg mL−1 and a low limit of detection (1.6 pg mL−1) to be obtained. In addition, a good selectivity against other non-target proteins was achieved. The developed method was validated by analyzing a WHO 1st International Standard for IFN-γ. In addition, the immunosensor was used for the determination of the endogenous IFN-γ in saliva with results in excellent agreement with those obtained by a commercial ELISA kit.

Enzymatic deposition of gold nanoparticles at vertically aligned carbon nanotubes for electrochemical stripping analysis and ultrasensitive immunosensing of carcinoembryonic antigen.

Herein we combine the sandwich immunoreaction at a vertically aligned single-walled carbon nanotube (SWCNT)-based immunosensor and the enzymatically catalytic deposition of gold nanoparticles (Au NPs) by a gold nanoprobe to develop a novel electrochemical immunosensing method. The vertically arranged nanostructure was prepared through the covalent linking of terminally carboxylated SWCNTs at an aryldiazonium-modified electrode. It not only provides an excellent platform for the high density immobilization of antibodies to obtain the immunosensor but also serves as useful molecular wires to accelerate electron transfer during the electrochemical immunosensing process. Meanwhile, the enzymatic reaction of the nanoprobe prepared by surface functionalization of the nanocarrier of Au NPs by high-content glucoamylases can catalyze the deposition of a large number of Au NPs at the immunosensor. The electrochemical stripping analysis of these nanoparticles enabled the convenient signal transduction of the method. Due to the sensitive gold stripping analysis at the vertically aligned SWCNTs and the multi-enzyme signal amplification of the nanoprobe, the electrochemical signal response was greatly enhanced. Thus, the method can be used for the ultrasensitive detection of the tumor biomarker of carcinoembryonic antigen in a wide linear range of 5 orders of magnitude with a low detection limit of 0.48 pg mL-1. Considering its obvious performance superiorities, this immunosensing method exhibits an extensive prospect for practical applications.

Resonance Raman scattering-infrared absorption dual-mode immunosensing for carcinoembryonic antigen based on ZnO@SiO2 nanocomposites

Detection of cancer biomarkers is crucial for the diagnosis and monitoring of malignant tumors. However, the accuracy and sensitivity still require sufficient improvement for practically clinical application. In this work, a reliable and sensitive dual-mode immunosensing method is described for carcinoembryonic antigen (CEA) detection using a biofunctional ZnO@SiO2 nanocomposite as a resonance Raman scattering (RRS)-infrared (IR) absorption nanoprobe. The multiphonon RRS signal originating from the ZnO and the characteristic IR fingerprint signal of the transverse optical and longitudinal optical phonon modes of the asymmetric stretching of Si–O–Si bonds showed no interference with each other. A CEA antibodies-immobilized substrate was fabricated to capture the analyte/nanoprobe complexes. The RRS intensity at 569 cm‒1 and the IR absorption at 1061 cm‒1 were used for quantitative analysis. Accurate CEA detection was performed as a result of the strong resistance of the dual-mode nanoprobe to surrounding interference. The limit of detection was 98.0 fg mL‒1. The detection range was 500 ng mL‒1 – 50 fg mL‒1, which is wider than those of single-mode RRS or IR absorption immunosensings. High reproducibility, selectivity and specificity were achieved. The assay performance of human serum samples demonstrated the practicability of the method in clinical cancer diagnosis.