Sensor thrombin

Tung, C. H., Gerszten, R. E., Jaffer, F. A. and Weissleder, R. (2002). A novel near-infrared fluorescence sensor for detection of thrombin activation in blood. Chembiochem 3, 207-211. 

Fig. 8.2 Nyquist plot of impedimetric sensor (Section 8.4.3.3). The redox couple ferro/ ferricyanide recorded at (a) bare Au electrode (b) Au electrode modified with thrombin-binding aptamer (c) Au/aptamer electrode with added 2-mercapto ethanol and (d) electrode with bound complementary thrombin/aptamer complex. Each plot was obtained separately (adapted from Radi etal.,2005)...

Low-wave acoustic sensor [63] was used to detect interaction of thrombin with RNA and DNA aptamers [24]. The authors compared the binding of thrombin to RNA aptamer also by using filter binding method utilizing radiolabeling of RNA aptamer by 3 -P32 and also by... 

An example of the DPV for thrombin DNA sensor is shown in Fig. 47.2. By integration of DPV using for example Origin 5.0 or higher version it is possible to determine total charge transfer for each record and to construct calibration curve as relative changes of charge transfer... 

Fig. 47.3. The plot of the relative changes of charge consumption during reduction of MB as a function of the concentration of the thrombin for two independently prepared sensors.

Fig. 3 Electrochemical aptamer-based sensor of redox-tagged DNA against specific targets, (a) When the aptamer comes in contact with a small molecule, in this case cocaine, it folds, and the redox tag is brought closer to the electrode, increasing the current, (b) When the aptamer comes in contact with thrombin, the tag moves away from the surface, decreasing the electrochemical signal. Reproduced from [85] with permission. Copyright Langmuir, 2007...

Fig. 3.2 Model illustrating the TBA aptamer modified sensor surface before and after incubation with Thrombin [49]...

Fig. 3.7 Spectral changes caused by thrombin (10 nM) binding to NSTBA sensor. The draninant peaks can be assigned to DNA vibrational modes with a lesser contribution from protein vibrational modes. Controls with BSA (10 mg/ml), msulin 600 nM and 1% human serum show no nonspecific protein binding. Lower three traces show spectra of NS, NS plus blocking agent (mercaptohexanol) and the fully assembled NSTBA sensor. Spectra are averages of 10 acquisitions...

To study the reproducibility and the reusability of the sensor, successive washing steps were performed with solutions of 8 M Urea and 0.2 M Ca EDTA followed by re-incubation with 10 nM thrombin. Urea was chosen to destabilize the protein aptamer complex and Ca /EDTA was chosen to disrupt the G-quadruplex. Spectra recorded after washing exhibit the low intensity unbound signal and signal increases repeatedly after incubation with 10 nM thrombin - these two states can be... 

Fig. 3.9 Washing and target incubation cycles of NSTBA with 10 nM thrombin. Graph shows intensities of dominant peaks. Peak intensities are highly reproducibly at every incubation cycle. A further incubation in which 10 nM thrombin was dissolved in a 1% solution of human serum produced similar peak intensities. Control experiments incubating only with bovine serum albumin, insulin, and human serum do not show any increased signals compared to the washed sensor [49]... 

Fig. 3.10 Result of the limit of detection (LOD) determination for NS-TBA in binding buffer for the cognate target protein human a thrombin. The sensor was incubated with protein concentrations between 500 yM and 50 pM. The red line indicates the LOD (mean plus 3 standard deviations of the blank measurement) [49]...

Zhu H, Suter JD, White IM, Fan X (2006) Aptamer based microsphere biosensor for thrombin detection. Sensors 6 785-795... 

To check the specificity of the sensor HSA at high concentration (77 pM), in a 1,400-fold excess respect to thrombin, is used. The interaction did not result in a measurable frequency decrease (AF < 3 Hz), demonstrating the high specificity of the sensor. 

In the past few years a new direction in TBA-based biosensors has been the development of methods for the electrochemical detection of thrombin. Some of the reported advantages of these electrochemical biosensors are their potential to provide high sensitivity, fast response times, low costs, easy fabrication, and the possibility for miniaturization. Ikebukoro and colleagues were the first to report on a TBA-based electrochemical sensor. Like others. 

Aptamer-based protein sensors have been developed since 1998. Thus, 0.7 amol of thrombin in 140-pL sample (0.5 pM concentration) was detected in a single-aptamer sensor through binding to a fluorescently labeled DNA aptamer by evanescent-wave-induced fluorescence anisotropy in less than 10 minutes [45]. Performance of DNA... 

Efficient immobilization of aptamers on surfaces is necessary for the construction of tongh, stable sensors and assay systems as one necessary step to overcome limitations for practical applications (Bini et al., 2007). Bini et al. (2007) have compared thrombin aptamers immobilized on a gold snrface by chemisorption (thiolated aptamer) and by biotin-streptavidin interaction (biotinylated aptamer carrying a linker) on a gold surface modified by a thiol-dextran-streptavidin layer. The linker-modified aptamer immobilized via streptavidin-biotin showed better reproducibility and sensitivity results for the quartz crystal sensor. Aptamers can be used for the functionalization of titanium-alloy surfaces (e.g., implant material, scaffolds) to enhance cell adhesion. The aptamers directed to osteoblasts are fixed electrochemically on the snrface of the alloy and promote cell adhesion (Gno et al., 2005, 2007). 

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