Aptasensor For Thrombin Detection Research Articles

PEC thrombin aptasensor based on Ag–Ag2S decorated hematite photoanode with signal-down effect of precipitation catalyzed by G-quadruplexes/hemin

A photoelectrochemical (PEC) aptasensor for thrombin detection was rationally designed based on the photoanode of one-dimensional hematite nanorods (α-Fe2O3 NRs) with several steps of modifications. Uniform α-Fe2O3 NRs were grown vertically on the surface of fluorine-doped tin oxide (FTO) conductive glass through a one-step hydrothermal method; then Ag was grown on the surface of α-Fe2O3 NRs through a photoreduction method followed by a partial in-situ transformation into Ag2S, conferring an improvement on the initial photocurrent. Two main critical factors, namely, the steric hindrance of thrombin, benzoquinone (BQ) precipitation oxidized by H2O2 under the catalysis of G-quadruplexes/hemin, contributed to the sensitive signal-down response toward the target. Photocurrent signals related with thrombin concentration was established for thrombin analysis due to the non-conductive complex as well as their competitive consumption of electron donors and irradiation light. The excellent initial photocurrent was combined with the signal-down amplification in the design of the biosensor, conferring a limit of detection (LOD) as low as 40.2 fM and a wide linear range from 0.0001 nM to 50 nM for the detection of thrombin. The proposed biosensor was also assessed in terms of selectivity, stability, and applicability in human serum analyses, which provided an appealing maneuver for the specific analysis of thrombin in trace amount.

An Enzyme- and Label-Free Fluorescence Aptasensor for Detection of Thrombin Based on Graphene Oxide and G-Quadruplex.

An enzyme- and label-free aptamer-based assay is described for the determination of thrombin. A DNA strand (S) consisting of two parts was designed, where the first (Sa) is the thrombin-binding aptamer and the second (Se) is a G-quadruplex. In the absence of thrombin, Sa is readily adsorbed by graphene oxide (GO), which has a preference for ss-DNA rather than for ds-DNA. Upon the addition of the N-methyl-mesoporphyrin IX (NMM), its fluorescence (with excitation/emission at 399/610 nm) is quenched by GO. In contrast, in the presence of thrombin, the aptamer will bind thrombin, and thus, be separated from GO. As a result, fluorescence will be enhanced. The increase is linear in the 0.37 nM to 50 µM thrombin concentration range, and the detection limit is 0.37 nM. The method is highly selective over other proteins, cost-effective, and simple. In our perception, it represents a universal detection scheme that may be applied to other targets according to the proper choice of the aptamer sequence and formation of a suitable aptamer-target pair.

L-cysteine induced manganese porphyrin electrocatalytic amplification with 3D DNA-Au@Pt nanoparticles as nanocarriers for sensitive electrochemical aptasensor

We successfully applied the novel l-cysteine (RSH) induced manganese porphyrin (MnTPP) electrocatalytic amplification in electrochemical aptasensor for detection of thrombin (TB) with the 3D DNA-Au@Pt nanoparticles (DNA-Au@Pt NPs) as nanocarrier. A further discussion for the suggested mechanism of the reaction involved the MnTPP-catalyzed oxidation of thiols (RSH) to disulfides l-cystine (RSSR) was also discussed in detail in this work. In comparison with traditional H2O2 mediated MnTPP electrocatalytic amplification, the novel reaction with more stable catalytic substrate RSH possessed high catalytic amplification efficiency. In addition, the 3D DNA-Au@Pt NPs could provide abundant binding sites for immobilizing signal tags and enzymes. As a result, the electrochemical signal can be greatly enhanced by the RSH mediated MnTPP electrocatalytic reaction and 3D DNA-Au@Pt NPs. Under optimal conditions, the proposed aptasensor exhibited a wider linear range of 0.1pM–100nM with a more sensitive detection limit of 29fM for TB detection.

An Improved Turn‐On Aptasensor for Thrombin Detection Using Split Aptamer Fragments and Graphene Oxide

AbstractAn improved turn‐on aptasensor for thrombin detection using split aptamer fragments and graphene oxide (GO) was reported. The thrombin‐binding aptamer (Apt15) was split into two parts for target recognition, an 8‐base sequence labeled with fluorescein (FAM‐Apt‐A) and a 7‐base oligonucleotide sequence (Apt‐B). In the absence of target protein, the fluorescence of FAM‐Apt‐A/Apt‐B was quenched by GO through Π‐Π stacking between GO and single‐stranded DNA. However, when thrombin was introduced into the system, a target‐induced G‐quadruplex forms with two split aptamer fragments and thrombin. The fluorescence recovered due to weak interaction between G‐quadruplex and GO. Compared to the strategy using intact aptamer, probe concentration was lowered, and an improved sensitivity was obtained. Moreover, heating process to avoid unfavorable secondary structure was avoided due to the use of shorter split aptamer fragments.

An anti-fouling aptasensor for detection of thrombin by dual polarization interferometry.

An anti-fouling surface was designed to effectively resist nonspecific protein adsorption using dual polarization interferometry, based on which the aptasensor for detection of thrombin was fabricated according to the specific interaction between thrombin and its 15-mer aptamer.

A label-free and sensitive electrochemical aptasensor for thrombin based on the direct electron transfer of hemin and hemin@rGO nanosheets as the signal probe

In this work, a label-free electrochemical aptasensor for sensitive detection of thrombin was fabricated and characterized.

A New Electrochemical Aptasensor for Protein Detection Based on Target Protein-Induced Strand Displacement

A sensitive electrochemical aptasensor for detection of thrombin based on target protein-induced strand displacement is presented. For this proposed aptasensor, dsDNA which is from immobilized probe DNA (IP) which has a hybridized with aptamer(Apt) initially was immobilized on the Au electrode through thiol group of IP，while a DNA labeled with PdS nanoparticles was used as a detection probe (DP-PdS). When Au electrode immersed into solution which contain target protein and DP-PdS, the aptamer preferred to form G-quarter structure with the present target protein and dsDNA was released into immobilized probe DNA (IP) which could be hybridized with DP-PdS. After dissolving PdS from electrode, a mercury-film electrode was used for electrochemical detection of these Pd2+ ions which offered excellent electrochemical signal transduction. Detection of thrombin was specific without being affected by the coexistence of other proteins such as BSA and lysozyme, and as low as 2.3×10-11 mol/L of thrombin was detected.

A sensitive and selective thrombin impedimetric aptasensor based on tailored aptamers obtained by solid-phase synthesis

The aim of the present work is to show the interest of a generic concept of aptamers directly obtained through a supported synthesis, using phosphoramidite synthons, for biosensor application. By that way, we were able to tailor the anchoring groups and specific spacers. The concept is validated on an impedimetric aptasensor for detection of thrombin. Two aptamer sequences, selected from literature (15 and 29 mers), were synthesized with a multi-thiol modification at one extremity. After their grafting on gold electrode, the faradic electrochemical impedance measurements allow the direct detection of thrombin in the range of 3–30ngmL−1and 3–50ngmL−1for 15 meraptamer and 29 meraptamer respectively, with a detection limit of 3.1ngmL−1 (80pmol/l) for both sequences. This study demonstrates that these aptasensors exhibit high sensitivity (higher sensitivity for 29 meraptamer), high selectivity (compared to a model protein, BSA), stability and reproducibility.

An Electrochemical Aptasensor for Detection of Thrombin based on Target Protein‐induced Strand Displacement

AbstractA sensitive electrochemical aptasensor for detection of thrombin based on target protein‐induced strand displacement is presented. For this proposed aptasensor, dsDNA which was prepared by the hybridization reaction of the immobilized probe ssDNA (IP) containing thiol group and thrombin aptamer base sequence was initially immobilized on the Au electrode by self‐assembling via AuS bind, and a single DNA labeled with CdS nanoparticles (DP‐CdS) was used as a detection probe. When the so prepared dsDNA modified Au electrode was immersed into a solution containing target protein and DP‐CdS, the aptamer in the dsDNA preferred to form G‐quarter structure with the present target protein resulting that the dsDNA sequence released one single strand and returned to IP strand which consequently hybridized with DP‐CdS. After dissolving the captured CdS particles from the electrode, a mercury‐film electrode was used for electrochemical detection of these Cd2+ ions which offered sensitive electrochemical signal transduction. The peak current of Cd2+ ions had a good linear relationship with the thrombin concentration in the range of 2.3×10−9–2.3×10−12 mol/L and the detection limit was 4.3×10−13 mol/L of thrombin. The detection was also specific for thrombin without being affected by the coexistence of other proteins, such as BSA and lysozyme.

A electrochemiluminescence aptasensor for detection of thrombin incorporating the capture aptamer labeled with gold nanoparticles immobilized onto the thio-silanized ITO electrode

A novel electrochemiluminescence (ECL) aptasensor was proposed for sensitive and cost-effective detection of the target thrombin adopted an aptamer-based sandwich format. To detect thrombin, capture aptamers labeled with gold nanoparticles (AuNPs) were first immobilized onto the thio-silanized ITO electrode surface through strong Au–S bonds. After catching the target thrombin, signal aptamers tagged with ECL labels were attached to the assembled electrode surface. As a result, an AuNPs-capture-aptamer/thrombin/ECL-tagged-signal-aptamer sandwich type was formed. Treating the resulting electrode surface with tri- n-propylamine (TPA) and applying a swept potential to the electrode, ECL response was generated which realized the detection of target protein. Spectroscopy and electrochemical impedance techniques were used to characterize and confirm the fabrication of the ECL aptasensor. AuNPs amplification and smart sensor fabrication art were implemented for the sensitive and cost-effective detection purpose. Signal-to-dose curve excellently followed a sandwich format equation and could be used to quantify the protein, and the detection limit was estimated to be 10 nM. Other forms of thrombin such as β- and γ-thrombins had negligible response, which indicated a high specificity of α-thrombin detection. The aptasensor opened up new fields of aptamer applications in ECL domain, a highly sensitive technique, and had a promising perspective to be applied in microarray analysis.

A new amplification strategy for ultrasensitive electrochemical aptasensor with network-like thiocyanuric acid/gold nanoparticles

An ultrasensitive and highly specific electrochemical aptasensor for detection of thrombin based on gold nanoparticles and thiocyanuric acid is presented. For this proposed aptasensor, aptamerI was immobilized on the magnetic nanoparticles, aptamerII was labeled with gold nanoparticles. The magnetic nanoparticle was used for separation and collection, and gold nanoparticle offered excellent electrochemical signal transduction. Through the specific recognition for thrombin, a sandwich format of magnetic nanoparticle/thrombin/gold nanoparticle was fabricated, and the signal amplification was further implemented by forming network-like thiocyanuric acid/gold nanoparticles. A significant sensitivity enhancement had been obtained, and the detection limit was down to 7.82aM. The presence of other proteins such as BSA and lysozyme did not affect the detection of thrombin, which indicates a high specificity of thrombin detection could be achieved. This electrochemical aptasensor is expected to have wide applications in protein monitoring and disease diagnosis.