Specific Detection Of Thrombin Research Articles

Hemin/G-quadruplex and AuNPs-MoS2 based novel dual signal amplification strategy for ultrasensitively sandwich-type electrochemical thrombin aptasensor

In this work, a novel sandwich-type electrochemical aptasensor based on the dual signal amplification strategy of hemin/G-quadruplex and AuNPs-MoS2 was designed and constructed, which realized the highly sensitive and specific detection of thrombin (TB). In this aptasensor, the 15-mer TB-binding aptamer (TBA-1) modified with thiol group was immobilized on the surface of AuNPs modified glassy carbon electrode (AuNPs/GCE) as capturing elements. Another thiol-modified 29-mer TB-binding aptamer (TBA-2) sequence containing G-quadruplex structure for hemin immobilization was designed. The formed hemin/G-quadruplex/TBA-2 sequence was further combined to the AuNPs decorated flower-like molybdenum disulfide (AuNPs-MoS2) composite surface via Au-S bonds, acting the role of reporter probe. In presence of the target TB, the sandwich-type electrochemical aptamer detection system could be formed properly. With the assistance of the dual signal amplification of AuNPs-MoS2 and hemin/G-quadruplex toward H2O2 reduction, the sandwich-type electrochemical aptasensor was successfully constructed for sensitive detection of TB. The results demonstrate that the fabricated aptasensor displays a wide linear range of 1.0 × 10−6 ∼ 10.0 nM with a low detection limit of 0.34 fM. This proposed aptasensor shows potential application in the detection of TB content in real biological samples with high sensitivity, selectivity, and reliability.

A novel LoC-SERS device integrated with aptamer recognition strategy for the highly sensitive and specific detection of thrombin and platelet-derived growth factor-B

A LoC-SERS device integrated with aptamer recognition strategy for the highly sensitive and specific detection of thrombin and platelet-derived growth factor-B.

A novel strategy for sensitive detection of thrombin via subtly integrated polypeptide substrate and aggregation-induced emission fluorophores in carotid artery thrombosis

Accurate monitoring thrombin played an important role in the diagnosis of carotid artery thrombosis. In this investigation, a novel strategy was designed by one-step process for sensitive and specific detection of thrombin via integrated 2-(4-bromophenyl)− 3-(4-(4-(diphenylamino)styryl) phenyl) fumaronitrile (TPABDFN) and polypeptide substrate. More concretely, TPABDFN was gathered and wrapped by the peculiar short chain polypeptide substrate (SPS) with the assistance of glutaric dialdehyde in water containing 10 % tetrahydrofuran (THF), possessing an “ON” state owing to the aggregation-induced emission (AIE) characteristics. In the present of thrombin, the TPABDFN-SPS could be specifically degraded, leading the “OFF” state by the specific hydrolysis of thrombin. Indeed, the fluorescence intensity of the TPABDFN-SPS would decline with the increasing concentrations of thrombin. The results demonstrated that this “turn-off” fluorescent complex had a linear response in the range of 1.56–200 ng mL−1 for thrombin with the detection limit of 0.42 ng mL−1. More fascinatingly, this strategy could further applied to monitor thrombin in the plasma sample, which displayed that the thrombin concentration in the plasma of carotid artery thrombosis (CAT) rats was higher than that in the sham group. Conceivably, the proposed strategy showed prospective promise in the detection of target proteases for the diagnose of certain disease.

Recent Progresses in Development of Biosensors for Thrombin Detection.

Thrombin is a serine protease with an essential role in homeostasis and blood coagulation. During vascular injuries, thrombin is generated from prothrombin, a plasma protein, to polymerize fibrinogen molecules into fibrin filaments. Moreover, thrombin is a potent stimulant for platelet activation, which causes blood clots to prevent bleeding. The rapid and sensitive detection of thrombin is important in biological analysis and clinical diagnosis. Hence, various biosensors for thrombin measurement have been developed. Biosensors are devices that produce a quantifiable signal from biological interactions in proportion to the concentration of a target analyte. An aptasensor is a biosensor in which a DNA or RNA aptamer has been used as a biological recognition element and can identify target molecules with a high degree of sensitivity and affinity. Designed biosensors could provide effective methods for the highly selective and specific detection of thrombin. This review has attempted to provide an update of the various biosensors proposed in the literature, which have been designed for thrombin detection. According to their various transducers, the constructions and compositions, the performance, benefits, and restrictions of each are summarized and compared.

Label-Free Quartz Crystal Microbalance Biosensor Based on Aptamer-Capped Gold Nanocages Loaded with Polyamidoamine for Thrombin Detection

Thrombin is an important biomarker, and its detection is of great significance for diagnosis and prevention of related diseases. Conventional methods such as enzyme-linked immunosorbent assay are sensitive but require multiple steps. In this work, a label-free biosensor that only required one step of incubation was developed based on target-triggered release of the cargo molecules from gold nanocages, which achieved highly sensitive and specific detection of thrombin. The proposed biosensor consists of an array of gold nanocages loaded with molecules in their interiors and the DNA probes immobilized on their surface for hybridization with thrombin-specific aptamers to seal their pores. Upon interaction with thrombin, the surface aptamers were lifted off the gold nanocages, resulting in release of the cargo molecules. The loss of cargo molecules was detected by quartz crystal microbalance (QCM). The signal could be amplified by the choice of cargo molecules. The use of polyamidoamine as cargo molecules allowed us to achieve a label-free biosensor with a linear detection range of 0.0086–86 nM and a limit of detection of 7.7 pM. The biosensor was also tested with spiked human serum samples with a limit of detection of 1.2 nM. The detection could be carried out within 1.5 h with only one incubation step of 45 min. The specificity of the biosensor was confirmed by testing against bovine serum albumin and lysozyme at 1 μM.

Dual-recognition colorimetric sensing of thrombin based on surface-imprinted aptamer-Fe3O4.

Thrombin plays an essential role in blood coagulation and some physiological and pathological processes. The convenient, rapid, sensitive, and specific detection of thrombin is of great significance in clinical research and diagnosis. Herein, surface molecularly imprinted polymer (MIP) was modified on aptamer-functionalized Fe3O4 nanoparticles (MIP-aptamer-Fe3O4 NP) for thrombin colorimetric assay by taking advantage of the peroxidase-like activity of Fe3O4 NP. With the adsorption of thrombin into imprinted cavities, the exposed surface area of Fe3O4 NP decreased, causing a decrease in its peroxidase-like activity toward 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of H2O2. On the other hand, the reductive amino acids on the thrombin surface also impeded the oxidation of TMB. Both phenomena caused the light blue color of the sensing solution. Thus, a specifically sensitive colorimetric approach for the visual detection of thrombin was proposed with a linear range and limit of detection of 108.1 pmol L-1-2.7 × 10-5 mol L-1 and 27.8 pmol L-1, respectively. Moreover, due to the double recognition elements of MIP and aptamer, the prepared MIP-aptamer-Fe3O4 NP showed higher selectivity to thrombin than that based on only one recognition element. It is worth noting that no special property (e.g. electrochemical or fluorescence activity) of the template was required in this work. Thus, more template molecules can be easily, selectively, and sensitively detected based on the proposed MIP-aptamer-mimic enzyme colorimetric sensing strategy.

A photoelectrochemical aptasensor for thrombin based on the use of carbon quantum dot-sensitized TiO2 and visible-light photoelectrochemical activity.

A photoelectrochemical (PEC) aptasensor for the highly sensitive and specific detection of thrombin is described. This aptasensor is based on an indium tin oxide (ITO) support that is covered with carbon quantum dot (CQD)-sensitized TiO2 and acts as a photoactive matrix. The ITO/TiO2/CQD electrode was prepared by impregnation assembly. Itdisplays an enhanced and steady photocurrent response under irradiation by visible light. A carboxyl-functionalized thrombin-binding aptamer was covalently immobilized on the modified ITO to obtain a PEC aptasensor whose photocurrent decreases with increasing concentration of thrombin. Under 420nm irradiation at a bias voltage of 0V, the aptasensor has a linear response in the 1.0 to 250 pM thrombin concentration range,with a 0.83 pM detection limit. Conceivably, this approach can be extended to numerous other PEC aptasensors for the detection of targets for which appropriate aptamers are available. Graphical abstract Schematic of a PEC aptasensor for thrombin. It is based on the use of CQD as the sensitizer, TiO2/CQDs as the photoactive matrix, and the thrombin aptamer as the recognition element.

Multi-DNAzymes-functionalized gold nanoparticles for ultrasensitive chemiluminescence detection of thrombin on microchip

Herein, a chemiluminescence assay with dual signal amplification has been developed based on multi-DNAzymes-functionalized gold nanoparticles (AuNPs) using in situ rolling circle amplification (RCA) for ultrasensitive detection of thrombin on microchip. In this assay, AuNPs was functionalized by aptamer and multi-RCA primer for amplification, and thrombin was sandwiched between the aptamer modified on the microchannel and the aptamer linked AuNP. The further amplification was realized by in situ RCA to expand specific oligonucleotides chains on the AuNPs and produce particular multi-DNAzymes. Enhanced chemiluminescence signal was achieved by the catalytic effect of DNAzymes in the luminol-H2O2 system. The sensitivity of detection was greatly improved by the dual amplification of multi-RCA primer modified AuNPs, and RCA. The whole strategy was applied for ultrasensitive and specific detection of thrombin. The chemiluminesce assay of thrombin performed a good linear range of 1–25 pM and the limit of detection was as low as 0.55 pM. The successful determination of thrombin in real human serum sample indicated a great potential in clinical study.

DNA-Mediated rolling circle amplification for ultrasensitive detection of thrombin using MALDI-TOF mass spectrometry.

A DNA-mediated rolling circle amplification (RCA) strategy was established for ultrasensitive and specific detection of thrombin via matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). This platform provides a versatile assay for the detection of various biomolecules of low abundance.

Construction of photoelectrochemical thrombin aptasensor via assembling multilayer of graphene–CdS nanocomposites

A photoelectrochemical (PEC) aptasensor for highly sensitive and specific detection of thrombin was developed by using graphene–CdS nanocomposites multilayer as photoactive species and electroactive mediator hexaammineruthenium(III) chloride (RuNH363+) as signal enhancer. Graphene–CdS nanocomposites (G–CdS) were synthesized by one-pot reduction of oxide graphene and CdCl2 with thioacetamide. The photoactive multilayer was prepared by alternative assembly of the negatively charged 3-mercaptopropionic acid modified graphene–CdS nanocomposites (MPA-G–CdS) and the positively charged polyethylenimine (PEI) on ITO electrode. This layer-by-layer assembly method enhanced the stability and homogeneity of the photocurrent readout of G–CdS. Thrombin aptamer was covalently bound to the multilayer by using glutaraldehyde as cross-linking. Electroactive mediator RuNH363+ could interact with the DNA phosphate backbone and thus facilitated the electron transfer between G–CdS multilayer and electrode and enhanced the photocurrent. Hybridizing of a long complementary DNA with thrombin aptamer could increase the adsorption amount of RuNH363+, which in turn boosted the signal readout. In the presence of target thrombin, the affinity interaction between thrombin and its aptamer resulted in the long complementary DNA releasing from the G–CdS multilayer and decreasing of photocurrent signal. On the basis of G–CdS multilayer as the photoactive species, RuNH363+ as an electroactive mediator, and aptamer as a recognition module, a high sensitive PEC aptasensor for thrombin detection was proposed. The thrombin aptasensor displayed a linear range from 2.0pM to 600.0pM and a detection limit of 1.0pM. The present strategy provided a promising ideology for the future development of PEC biosensor.

Assembly of a Dual Aptamer Gold Nanoparticle Conjugate Ensemble in the Specific Detection of Thrombin when Coupled with Dynamic Light Scattering Spectroscopy

We demonstrate an extremely facile, rapid, specific and selective method for detecting proteins using aptamerconjugated gold nanoparticles coupled with dynamic light scattering (DLS) at ambient conditions. Addition of proteins to aptamer-conjugated gold nanoparticles (GNPs) induced the formation of protein-aptamer-gold nanoparticle conjugate complexes. The average hydrodynamic diameter of the nanoconjugate complexes as measured by DLS, increased with the corresponding increase in protein concentration. This correlation formed the analytical basis of the assay. The nanoparticles and nanoconjugate complexes were characterized by transmission electron microscopy, ultra-violet visible spectroscopy and DLS. Various parameters affecting the assay were optimized. Using thrombin as the model analyte, we demonstrated the detection of as low as 1.41 nM (0.05 μg/mL) of the protein. A linear dynamic range of up to 300 nM (11 μg/mL) was realized. The presence of other interfering proteins such as BSA showed no effect on the assay response.The presence of other interfering proteins such as bovine serum albumin (BSA) showed no significant effect on the assay response.