Aptamer Sensor Research Articles

Development of a SERS aptasensor for the determination of L-theanine using a noble metal nanoparticle-magnetic nanospheres composite.

An ultrasensitive surface-enhanced Raman spectroscopy (SERS) aptamer sensor (aptasensor) using a noble metal nanoparticle-magnetic nanospheres composite was developed for L-theanine detection. It makes use of Fe3O4@Au MNPs and Au@Ag NPs embedded with the Raman reporter 4-mercaptobenzoic acid (4MBA). Au@4MBA@Ag NPs modified by aptamer and Fe3O4@Au MNPs modified by cDNA created the aptasensor with the strongest Raman signal of 4MBA through the specific binding of the aptamer. With the preferred binding of L-theanine aptamer to L-theanine, Au@4MBA@Ag NPs were released from Fe3O4@Au MNPs, causing a linear decrease in SERS intensity to achieve the SERS detection of the L-theanine. The SERS peak of 4MBA at 1078cm-1 was used for quantitative determination. SERS intensity showed a good log-linear relationship within the range 10-10 to 10-6Mof L-theanine. Theaptasensor has a high selectivityfor L-theanine compared with other twelve testedanalytes. Hence, thisaptasensor is a promising analytical tool for L-theanine detection. The developedmethod was appliedto the analysis of real samples, demonstrating excellent performance. The comparison with the standard liquid chromatography mass spectrometry methodshowed an error within 20%.

Construction of an aptamer sensor for sensitive detection of AOH based on the enhancement of light/electric dual signaling response of Ru/Cu-THQ by Ag NCs

Precise detection of food safety is a hotspot and challenging aspect in contemporary scientific research. This study utilized Ag nanocubes (Ag NCs) to augment the electrogenerated chemiluminescence (ECL) and electrochemical (EC) dual-response signals of Ru/Cu-tetrahydroxybenzoquinone (Ru/Cu-THQ). Consequently, a novel aptamer sensor was developed for the trace detection of Alternariol (AOH). This dual-mode sensor offers several advantages: (1) The output of dual-channel signals significantly reduces the likelihood of false-positive and false-negative detection results, which are common in traditional single-mode sensing. (2) Ru/Cu-THQ effectively coordinates dual-channel outputs and exhibits excellent ECL and EC behavior. (3) Ag NCs, due to their strong K2S2O8 activation ability and excellent conductivity, further amplify the sensor's response signal. This sensor demonstrates effective detection performance for AOH in the concentration range of 0.01–100 ng/mL, offering a novel approach for accurate food safety detection and potential application in real sample analysis.

Dual-mode colorimetric electrochemical aptamer sensor based on Au@Pd bimetallic nanoparticles for the detection of Pb(II) in edible mushrooms

Dual-mode colorimetric electrochemical aptamer sensor based on Au@Pd bimetallic nanoparticles for the detection of Pb(II) in edible mushrooms

Aptamer-based fluorescent sensor for highly sensitive detection of methamphetamine.

The construction of a fluorescence aptamer sensor was achieved by employing the fundamental principle of fluorescence resonance energy transfer. By employing molecular modeling technologies to identify the binding site, the high-affinity aptamer APT-40nt was derived from the whole sequence and utilized on the graphene oxide (GO) fluorescent platform for the purpose of achieving a highly sensitive detection of methamphetamine (METH). The aptamer tagged with fluorescein (FAM) dye undergoes quenching in the presence of GO due to π-stacking interaction. With the addition of the target, the aptamer that has been tagged was detached from the GO surface, forming a stable complex with METH. This process resulted in fluorescence restoration of the system, and the degree of fluorescence restoration was proportional to METH concentration in the linear range of 1-50 and 50-200 nM. Notably, under optimized conditions, the detection limit of this aptasensor was as low as 0.78 nM, which meets the detection limit requirements of METH detection in saliva and urine in some countries and regions. Moreover, other common illicit drugs and metabolites had minimizing interference with the determination. The established aptasensor, therefore, has been successfully applied to detect METH in saliva and urine samples and exhibited satisfactory recoveries (87%-111%). This aptasensor has the advantages of low detection limit, excellent selectivity, ease of operation, and low cost, providing a promising strategy for on-site detection of METH in saliva and urine.

A photoelectrochemical aptasensing platform assembled at the heterojunction interface of Cu3BiS3 sensitized CuV2O6 for bisphenol A.

The interaction between the sensitive interfaces of photoelectrochemical (PEC) semiconductor nanomaterials and microscopic matter creates endless potential for the efficient detection of endocrine disruptor. Thiswork presents the development of a high-efficiency PEC aptasensor for bisphenol A (BPA) monitoring based on Cu3BiS3 sensitized CuV2O6 nanocomposites with exceptional visible-light PEC activity. We implemented the integration of Cu3BiS3 nanosheet photosensitizer to sensitize the CuV2O6 nanowire structure that was synthesized utilizing a facile hydrothermal approach. The band gap alignment between Cu3BiS3 and CuV2O6 facilitated enduring PEC response yielding an efficient interfacial structure. The surface of the CuV2O6/Cu3BiS3 electrode was modified with BPA aptamer, enabling specific binding with BPA and precise quantification of its content. The developed aptamer sensors possess a wide detection range of5.00 × 10-1 to 5.00 × 104 ng/mL, and a low detection limit of 1.60 × 10-1 ng/mL (at S/N = 3). After undergoing 20 testing cycles and enduring long-term storage, the sensor maintained its stability and showcased excellent repeatability and reproducibility. This study presents a promising methodology for the detection of BPA in environmental settings.

A fluorescence aptamer sensor utilizing WS2 nanosheets for sensitive detection of patulin: enhanced specificity and wide applicability.

A tungsten disulfide (WS2) nanosheet-based aptamer sensor was developed to detect patulin (PAT). The 5'-end of the PAT aptamer was modified with a cyanine 3 (Cy3) fluorophore, which self-assembled on WS2 nanosheets. The interaction between the Cy3 fluorophore at the 5'-end of the PAT aptamer and the WS2 nanosheets resulted in reduced fluorescence (FL) intensity due to fluorescence resonance energy transfer (FRET). The introduction of PAT into this sensing system led to hybridization with the PAT aptamer, forming a G-quadruplex/PAT complex with low affinity for the WS2 nanosheet surface. This hybridization increased the distance between the Cy3 fluorophore and the WS2 nanosheets, inhibiting FRET and producing a strong FL signal. Under optimal experimental conditions, the FL intensity of the sensing system demonstrated an excellent linear correlation with PAT concentrations ranging from 0.5 to 40.0 ng mL-1, and it achieved a detection limit (S/N = 3) of 0.23 ng mL-1. This sensing system offers enhanced specificity for PAT detection and has the potential for broad application in detecting other toxins by substituting the sequence of the recognition aptamer.

Electrochemical aptamer sensor based on AgNPs@PDANSs and "sandwich" structure guidance for the detection of tobramycin in water samples.

In this study, an ultrasensitive detection platform for tobramycin (TOB) was developed, featuring a "sandwich" structure guided by AgNCs@PDANSs and Thi-AuNCs@ZnONSs. To address the issue of large background current peak signals in tagless sensors, Thi-AuNCs@ZnONSs composites were synthesized as signal tags. Zinc oxide nanosheets (ZnONSs) served as the loading agent, and AuNCs with the electroactive molecule Thi acted as carriers. Furthermore, AgNPs@PDANSs nanocomposites, possessing excellent electrical conductivity and large specific surface areas, were prepared as substrate materials for the modified electrodes. A "sandwich" structure strategy was also introduced to enhance the accuracy of the electrochemical aptasensor. This strategy, utilizing a dual sequence for target labeling and capture, yielded higher sensitivity and simplified the sensor construction compared to methods employing a single sequence. Under optimal conditions, the detection limit for TOB was established at 1.41 pM, with a detection range of 0.05-5000 nM. The aptasensor was effectively applied in the detection of TOB in tap and lake water, demonstrating outstanding reproducibility, selectivity, and stability. These results may serve as a reference for environmental TOB detection.

Development of a highly sensitive ampicillin sensor utilizing functionalized aptamers.

To develop a sensitive and simple ampicillin (AMP) sensor for trace antibiotic residue detection, the influencing factors of the modification effect of nanogold-functionalized nucleic acid sequences (Adenine: A, Thymine: T) were comprehensively analyzed in this study, including the modification method, base length and type. It was found that under the same base concentration, longer chains are more likely to reach saturation than shorter chains; and when the base concentration and length are both the same, A exhibits a higher saturation modification level compared to T. Based on these research findings, a highly sensitive fluorescence aptamer sensor for detecting ampicillin was constructed using the optimized functionalized sequence (ployA6-aptamer) and experimental conditions (6 hours binding time between nucleic acid aptamer and complementary strand, pH 7 working solution, 20 minutes detection time) based on the principle of fluorescence resonance energy transfer. The sensor has a detection range of 0.18 ng ml-1 to 3.11 ng ml-1 for ampicillin, with a detection limit of 0.04 ng ml-1. It exhibits significant selectivity and achieves an average recovery rate of 98.71% in tap water and 91.83% in milk. This method can be used not only for residual ampicillin detection, but also for highly sensitive detection of various antibiotics and small biological molecules by replacing the aptamer type. It provides a research basis for the design of highly sensitive fluorescence aptamer sensors and further applications of nanogold@DNA composite structures.

A novel photoelectrochemical aptasensor for alpha-fetoprotein assay based on a 3D self-supporting TiO2/CdS/BiOI Z-scheme heterojunction

Alpha-fetoprotein (AFP) is a characteristic marker of liver cancer. This study establishes a PEC aptamer sensor using BiOI/CdS/TiO2 heterojunction material and realizes accurate detection of AFP.

A label free fluorescent aptamer sensor based on the combined action of Graphene oxide and SYBR Green I for the detection of Aflatoxin B1.

Here, based on the characteristics of Graphene oxide(GO) and SYBR Green I(SGI) dye, an enzyme-free and label-free fluorescent biosensor with signal amplification through DNA strand reaction is proposed for the detection of Aflatoxin B1(AFB1) in food safety. Firstly, without the addition of AFB1, the substrate in the system includes a double stranded Apt-S with a long sticky end and two hairpins H1 and H2. Although the complementary pairing of bases may exhibit fluorescence due to the insertion of SGI dyes, the use of GO, which is highly capable of adsorbing single stranded parts and quenching fluorescence, cleverly reduces the background fluorescence. Adding the target AFB1 triggers DNA inter chain reactions, generating a large amount of long double stranded DNA H1-H2, thereby generating strong fluorescence signals under the action of SGI. More importantly, logical theory verification and computer simulation were conducted before biological experiments, providing a theoretical basis for the implementation of the biosensor. After analysis, the fluorescence biosensor exhibits a good linear relationship with AFB1 concentration in the range of 5-50nM, with a detection limit of 0.76nM. It also has good specificity, anti-interference ability, and practical application ability, and has broad application prospects in the field of food safety.

Ultra-sensitive detection of tumor necrosis factor alpha based on silver-coated gold core shell and magnetically separated recognition of SERS aptamer sensors.

A highly sensitive tumor necrosis factor α (TNF-α) detection method based on a surface-enhanced Raman scattering (SERS) magnetic patch sensor is reported. Magnetic beads (MNPs) and core shells were used as the capture matrix and signaling probe, respectively. For this purpose, antibodies were immobilized on the surface of magnetic beads, and then Au@4-MBN@Ag core-shell structures coupled with aptamers and TNF-α antigen were added sequentially to form a sandwich immune complex. Quantitative analysis was performed by monitoring changes in the characteristic SERS signal intensity of the Raman reporter molecule 4-MBN. The results showed that the limit of detection (LOD) of the proposed method was 4.37 × 10-15 mg·mL-1 with good linearity (R2 = 0.9918) over the concentration range 10-12 to 10-5 mg·mL-1. Excellent assay accuracy was also demonstrated, with recoveries in the range 102% to 114%. Since all reactions occur in solution and are separated by magnetic adsorption of magnetic beads, this SERS-based immunoassay technique solves the kinetic problems of limited diffusion and difficult separation on solid substrates. The method is therefore expected to be a good clinical tool for the diagnosis of the inflammatory biomarker THF-α and in vivo inflammation screening.

Aptamer Based SPREETA Sensor for the Detection of Porphyromonas gingivalis G-Protein.

We have developed an aptamer that specifically binds to Porphyromonas gingivalis to reduce the cellular damage caused by P. gingivalis infection and applied it as a biosensor. P. gingivalis is one of the major pathogens causing destructive periodontal disease among the periodontal microorganisms constituting complex biofilms. Porphyromonas gingivalis G-protein (PGP) known to play an important role in the transmission of germs was used as a target protein for the screening of aptamer. The aptamer that has binds to the G-protein of P. gingivalis, was screened and developed through the Systemic Evolution of Ligands by Exponential Energy (SELEX) method. Modified-Western blot analysis was performed with the aptamer which consisted of 38 single-stranded DNA to confirm the selectivity. ELONA (enzyme linked oligonucleotide assay) used to confirm that the aptamer was sensitive to PGP even at low concentration of 1 μg/ml. For the rapid detection of P. gingivalis, we constructed a surface plasmon resonance biosensor with SPREETA using the PGP aptamer. It was confirmed that PGP could be detected as low concentration as at 0.1 pM, which is the minimum concentration of aptamer sensor within 5 min. Based on these results, we have constructed a SPREETA biosensor based on aptamer that can bind to P. gingivalis G-protein. It can be used as an infection diagnosis system to rapidly diagnose and analyze oral diseases caused by P. gingivalis.

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

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.

Modular Engineering of Aptamer-Based Nanobiotechnology for Conditional Control of ATP Sensing.

Dynamic changes of intracellular, extracellular, and subcellular adenosine triphosphates (ATPs) have fundamental interdependence with the physio-pathological states of cells. Spatially selective in situ imaging of such ATP dynamics offers valuable mechanistic insights into the related biological activities. Despite significant advances in the design of aptamer sensors for ATP detection, the dearth of methods that enable precise ATP imaging in specific cellular locations remains a challenge in this field. This review focuses on the modular engineering of regulatable sensing technology via the integration of aptamer probe designs with advanced functional nanomaterials, allowing conditional control of ATP sensing and imaging with high spatial precision from subcellular organelles to living animals. Highlighting the recent advances in the design of photo-triggered nanosensors for spatiotemporally controlled ATP imaging, endogenously-triggered ATP sensing in a cell-selective manner, and spatially-controlled nanodevices for ATP imaging in specific organelles and extracellular microenvironments. Emphasis will be put on elucidating the principles of how nanotechnology can be applied to regulate the spatial precision of aptamer-based ATP sensing activities. The authors envision that this perspective provides insights into the engineering of aptamer-based nanobiotechnology for opening new frontiers in precise molecular sensing and other bio-applications.

An electrochemical aptamer sensor for rapid quantification sulfadoxine based on synergistic signal amplification of indole and MWCNTs and its electrooxidation mechanism

Unreasonable disposal of sulfadoxine (SDX) causing food safety and eventually result water pollution, and shows great significance of detection. A sensitive unit Aptamer (Apt) was adopted to construct an electrochemical sensor BSA/Apt/indole/MWCNTs/GCE with the synergistic amplification of multi-walled carbon nanotubes (MWCNTs) and indole for the detection of SDX. Optimization of the conditions allowed the detection of SDX shows a wide linear range from 0.10 to 1000.00 µmol/L by differential pulse voltammetry (DPV). The LOD and sensitivity were determined to be 0.033 µmol/L and 0.29 µA/(µmol/L·cm²), respectively. The recovery rate of SDX in pork and lake water samples range 98.57 %−104.93 %, 93.6 %−106.2 %, respectively. Both the relative standard deviation (RSD) was lower than 7.40 %, which proved that the sensor has good practicability. In addition, DFT calculation was established that a restricted of B3LYP and a basis set of 6–31 G, the electrooxidation of SDX involves two-electron and one-proton transfer.

A Novel Fluorescent Aptasensor Based on Mesoporous Silica Nanoparticles for Selective and Sensitive Detection of Saxitoxin in Shellfish

Background: Saxitoxin (STX) stands as one of the most potent marine biotoxins, exhibiting high lethality. Despite its severity, current treatments remain ineffective, and existing detection techniques are limited due to ethical concerns and technical constraints. Methods: Herein, an innovative approach was constructed for STX detection, utilizing mesoporous silica nanoparticles (MSN) as a foundation. This innovative, easy, and label-free aptamer (Apt)- sensor was fabricated. Apts were employed as molecular identification probes and "gated molecules," while rhodamine 6G was encapsulated within particles to serve as a signal probe. In a lack of STX, Apts immobilized on an MSN surface kept a "gate" closed, preventing signal probe leakage. Upon the presence of STX, the "gate" opened, allowing a particular binding of Apts to STX and a subsequent release of a signal probe. Results: Experimental results demonstrated a positive correlation between fluorescence intensity and concentrations of STX within a range of 1 to 80 nM, with an exceptional limit of detection of 0.12 nM. Furthermore, the selectivity and stability of a biosensor were rigorously evaluated, validating its reliability. Conclusion: This newly developed sensing strategy exhibits remarkable performance in STX detection. Its success holds significant promise for advancing portable STX detection equipment, thereby addressing a pressing need for efficient and ethical detection methods in combating marine biotoxin contamination.

Aptamer conjugated Si nanowire network/poly(pyrrole)-NTA structure for electrochemical sensing

The current study describes the design, analytical characterization, and testing of aptamer sensors based on Si nanonets (Si NN) for thrombin electrochemical detection. Si nanonets are thin sheet arrays of randomly aligned Si nanowires with controlled and programmable properties that may be transported and integrated on a variety of substrates. Si nanonets were placed onto a commercial glassy carbon working electrode to create our aptasensor. Then they were functionalized by conductive pyrrole polymerization, which was followed by aptamer bioreceptor grafting. Electrochemical detection of thrombin was carried out after optimizing the electrode properties. On the one hand, cyclic voltammetry analyses demonstrated a linear association between the acquired current and the logarithm of thrombin concentration from 5 nmol L−1 to 2 µmol L−1. Impedance measurements, on the other hand, revealed a linear relationship between the charge transfer resistance as well as the impedance modulus. Thus, for the first time, Si nanonet thin sheets were used as constitutive nanoporous parts of electrodes capable of detecting thrombin at concentration limits of 5 nM, considered as a risk factor or presence of thrombosis. This demonstrates the concept of using this type of nanomaterial to detect molecules of biological interest.

Dual-mode transfer response based on electrochemical and fluorescence signals for the detection of amyloid-beta oligomers (AβO).

An aptamer sensor has been developedutilizing a dual-mode and stimuli-responsive strategy for quantitative detection of AβO (amyloid-beta oligomers) through simultaneous electrochemical and fluorescence detection. To achieve this, we employed UIO-66-NH2 as a carrier container to load MB (Methylene Blue), and Fe3O4 MNPs (iron oxide magnetic nanoparticles) with aptamer (ssDNA-Fe3O4 MNPs) fixed on their surface for biological gating. The ssDNA-Fe3O4 MNPs were immobilized onto the surface of UIO-66-NH2 through hydrogen bonding between the aptamer and the -NH2 group on the surface of UIO-66-NH2, thereby encapsulating MB and forming ssDNA-Fe3O4@MB@UIO-66-NH2. During the detection of AβO, the aptamer selectively reacted with AβO to form the AβO-ssDNA-Fe3O4 complex, leading to its detachment from the surface of UIO-66-NH2. This detachment facilitated the release of MB, enabling its electrochemical detection. Simultaneously, the AβO-ssDNA-Fe3O4 complex was efficiently collected and separated using a magnet after leaving the container's surface. Furthermore, the addition of NaOH facilitated the disconnection of biotin modifications at the 3' end of the aptamer from the avidin modifications on the Fe3O4 MNPs. Consequently, the aptamer detached from the surface of Fe3O4 MNPs, resulting in the restoration of fluorescence intensity of FAM (fluorescein-5'-carboxamidite) modified at its 5' end for fluorescence detection. The dual-mode sensor exhibited significantly enhanced differential pulse voltammetry signals and fluorescence intensity compared to those in the absence of AβO. The sensor demonstrated a wide detection range of 10 fM to 10 μM, with a detection limit of 3.4 fM. It displayed excellent performance in detecting actual samples and holds promising prospects for early diagnosis of Alzheimer's disease.

A novel fluorescent sensor based on aptamer recognition and DNA walker amplification strategy and its determination of 17β-estradiol

17β-Estradiol (E2) is a common environmental endocrine and excessive estrogen will disturb the endocrine balance of human body and lead to diseases because of its accumulation in human body. The main aim of this study is to determine the trace amount of E2 in various foods. Based on aptamer specific recognition, DNA walker molecular machine and Exonuclease III-assisted dual cycle signal amplification technology, a fluorescent aptamer sensor was constructed and applied to the detection of 17β-estradiol. Atomic force microscopy (AFM) and polyacrylamide gel electrophoresis (PAGE) were used to verify the feasibility of DNA walker participating in the reaction. Under the optimal conditions, 17β-estradiol concentration showed a good linear relationship with fluorescence in the range of 5.0 × 10−10–1.8 × 10−8 M, and the detection limit was 5.6 × 10−11 M. The fluorescent aptamer sensor established in this study has good selectivity, simple operation and good specificity, and the results of its application in the determination of milk samples are satisfactory. Research findings revealed that the E2 concentration in water and milk is less than the safe limits, indicating the foods in the market are safe and can be consumed. Though the foods are safe in present content, the health risk still exists because of continuous intake of trace E2.

Label-Free Homogeneous Electrochemical Aptasensor Based on Size Exclusion/Charge-Selective Permeability of Nanochannel Arrays and 2D Nanorecognitive Probe for Sensitive Detection of Alpha-Fetoprotein.

The labeling-free and immobilization-free homogeneous aptamer sensor offers advantages including simple operation, low cost, and high sensitivity, demonstrating great potential in rapid detection of tumor biomarkers in biological samples. In this work, a labeling-free and immobilization-free homogeneous aptamer sensor was conveniently fabricated by combining size exclusion and charge-selective penetration of a nanochannel-modified electrode and two-dimensional (2D) nanorecognition probe which can realize selective and highly sensitive detection of alpha-fetoprotein (AFP) in serum. Vertically ordered mesoporous silica film (VMSF) with ultra-small, uniform, and vertically aligned nanochannels was easily grown on the simple, low-cost, and disposable indium tin oxide (ITO) electrode. Through π-π interaction and electrostatic force, the AFP aptamer (Apt) and electrochemical probe, tris(bipyridine)ruthenium(II) (Ru(bpy)32+), were coloaded onto graphene oxide (GO) through simple incubation, forming a 2D nanoscale recognition probe (Ru(bpy)32+/Apt@GO). Owing to the size exclusion effect of VMSF towards the 2D nanoscale probe, the electrochemical signal of Ru(bpy)32+/Apt@GO could not be detected. In the presence of AFP, the specific binding of AFP to the aptamer causes the dissociation of the aptamer and Ru(bpy)32+ from GO, resulting in their presence in the solution. The efficient electrostatic enrichment towards Ru(bpy)32+ by negatively charged VMSF allows for high electrochemical signals of free Ru(bpy)32+ in the solution. Linear determination of AFP ranged from 1 pg/mL to 1000 ng/mL and could be obtained with a low limit of detection (LOD, 0.8 pg/mL). The high specificity of the adapter endowed the constructed sensor with high selectivity. The fabricated probe can be applied in direct determination of AFP in serum.