Apt Sequence Research Articles

Recovery time reduction to decrease experimental duration (R2D2): A simple and universal method to accelerate NMR experiments

Acceleration techniques for one dimensional Nuclear Magnetic Resonance (1D NMR) are very useful, both for NMR enthusiasts and for chemists that use NMR for structural elucidation. To the latter, such techniques need to be straightforward. Recovery time Reduction to Decrease the experimental Duration (R2D2) relies on the incremental reduction of a pulse sequence's Recycle Time (TR). A pseudo-2D spectrum is acquired and after two Fourier transform, extraction and addition of the central rows, a 1D spectrum is obtained. Not only can it be applied to any pulse sequence that contains a TR, but it also requires only a list of recovery times and 2D processes to operate. With this method, we were able to easily reduce the experimental time by a factor of 2 and up to 4 using single-pulse, APT and DEPT 13C sequences. Moreover, R2D2 has the potential to be used on other low abundance nuclei (such as 15N or 2H) and numerous other pulse sequences.

Multimodal MRI for Estimating Her-2 Gene Expression in Endometrial Cancer.

To assess the value of multimodal MRI, including amide proton transfer-weighted imaging (APT), diffusion kurtosis imaging (DKI), and T2 mapping sequences for estimating human epidermal growth factor receptor-2 (Her-2) expression in patients with endometrial cancer (EC). A total of 54 patients with EC who underwent multimodal pelvic MRI followed by biopsy were retrospectively selected and divided into the Her-2 positive (n = 24) and Her-2 negative (n = 30) groups. Her-2 expression was confirmed by immunohistochemistry (IHC). Two observers measured APT, mean kurtosis (MK), mean diffusivity (MD), and T2 values for EC lesions. The Her-2 (+) group showed higher APT values and lower MD and T2 values than the Her-2 (-) group (all p < 0.05); there was no significant difference in MK values (p > 0.05). The area under the receiver operating characteristic curve (AUC) of APT, MD, T2, APT + T2, APT + MD, T2 + MD, and APT + MD + T2 models to identify the two groups of cases were 0.824, 0.695, 0.721, 0.824, 0.858, 0.782, and 0.860, respectively, and the diagnostic efficacy after combined APT + MD + T2 value was significantly higher than those of MD and T2 values individually (p = 0.018, 0.028); the diagnostic efficacy of the combination of APT + T2 values was significantly higher than that of T2 values separately (p = 0.028). Weak negative correlations were observed between APT and T2 values (r = -0.365, p = 0.007), moderate negative correlations between APT and MD values (r = -0.560, p < 0.001), and weak positive correlations between MD and T2 values (r = 0.336, p = 0.013). The APT values were independent predictors for assessing Her-2 expression in EC patients. The APT, DKI, and T2 mapping sequences can be used to preoperatively assess the Her-2 expression in EC, which can contribute to more precise treatment for clinical preoperative.

A signal-off aptasensor for the determination of Acinetobacter baumannii by using methylene blue as an electrochemical probe.

An electrochemical aptasensor has been developed to detect Acinetobacter baumannii (A. baumannii). The proposed system was developed by modifying carbon screen-printed electrodes (CSPEs) with a synthesized MWCNT@Fe3O4@SiO2-Cl nanocomposite and then binding A. baumannii-specific aptamer using covalent immobilization on the modified electrode surface and the interaction of methylene blue (MB) with Apt as an electrochemical redox indicator. As a result of the incubation of the A. baumannii bacteria as a target on the proposed aptasensor, a cathodic peak current density (Jpc) of MB decreased due to the formation of the Apt-A. baumannii complex and MB being released from the immobilized Apt on the surface of the modified electrode. In addition to increasing the electron transfer kinetics, the nanocomposite provides a relatively stable matrix to improve the loading Apt sequence. The suggested aptasensor was demonstrated to be capable of detecting A. baumannii with a linear range of 10.0-1.0 × 107 colony-forming unit (CFU) mL-1 and a detection limit of 1 CFU mL-1 (S/N = 3) using differential pulse voltammetry (DPV) studies at a working potential of ~0.29 V and a scan rate of 100 mV s-1. The outcomes revealed that the aptasensor exhibited high A. baumannii detection sensitivity, stability, reproducibility, and specificity.

An Appraisal of Aptasensors for Dopamine Measurement in Biofluids: A Review

Dopamine (DA) mediates various physiological functions across the body within the digestive and renal systems and in the blood circulation. It also plays a key role in motivated behaviors as a brain neurotransmitter (NT) in animals and humans and in the etiology of several neurodegenerative disorders such as Parkinson’s disease. Nevertheless, selective tracking of DA in biofluids in the presence of competitive chemicals and NTs remains a challenge. The aptamer (Apt)-based sensors (aptasensors) could offer a valid strategy for DA measurement in bodily fluids due to their high specificity. This study provides a comprehensive and comparative review of the state of the art of all the electrochemical and optical aptasensors for DA detection. The function of the aptasensors has been discussed in DA assay by considering various aspects such as the utilized Apt sequences, their immobilization protocol on the platform, engineered sensing interface, and diverse probes and modifiers in the electrochemical and optical approaches. Notably, a few of the published aptasensing techniques in both electrochemical or optical domains have already achieved very high sensitivity for DA tracking below picomolar and femtomolar range with temporal resolutions down to seconds. Resolution of some of remaining challenges, such as biocompatibility of substrates used and improving the temporal resolution, can enhance the prospects of using aptasensors in live tissue for chronic DA screening.

Electrochemical aptasensor based on the target-induced strand displacement strategy-driven for T-2 toxin detection

Herein, an aptasensor based on target-induced strand displacement (TISD) strategy was developed for sensitive detection of T-2 toxin. Gold nanoparticles@ aminated manganese dioxide (AuNPs@NH2-MnO2) exhibited excellent electrical conductivity and provided more binding sites for aptamer (Apt). Besides, polyethyleneimine-reduced graphene oxide/gold‑platinum core-shell nanorods composites (PEI-rGO/Pt@Au NRs) were used to be carriers for signaling tags, as their sufficiently large specific surface area improved the loading capacity for signal molecules. In the presence of T-2, the Apt sequence was more inclined to form an Apt-T-2 complex, and the cDNA was displaced from the Apt-cDNA duplex, while the signal tag was released, resulting in a weakened MB signal, differential pulse voltammetry (DPV) was used to record the signal change. Under optimal conditions, the signal response of the constructed electrochemical aptasensor exhibited a good linear relationship with the concentration of T-2. The detection limit was 8.74 × 10−7 ng mL−1over a wide range of concentration from 5 × 10−6 ng mL−1 to 5 ng mL−1. Furthermore, the proposed aptasensor had excellent specificity, good stability and can be well applied to the detection of real samples. It provided a new avenue for the research and development of sensitive aptasensors in food detection and analysis.

Development of a novel liquid crystal Apta-sensing platform using P-shape molecular switch

Described herein is a liquid crystal (LC)-based aptasensor via employing the reorientation of LC triggered by the conformational changes of a P-shaped DNA structure. The structure consists of a short linker sequence as an immobilizer probe with ability to hybridize with the central part of the intact aptamer (Apt) sequence and an Apt terminal-locker (ATL) strand with complementary segments of the Apt terminal fragments. Bindings of two arm segments of the Apt sequence with the ATL strand enforces it to form a P-shaped configuration on the sensing platform. The selective interaction between the Apt strand and OTA leads to the disassembly of the Apt-ATL hybrid, collapse of the P-shaped structure, and consequently, transition of the optical appearance of the aptasensor texture. Determination of Ochratoxin A (OTA) in foods is an urgent demand in attempt to minimize food safety risks. To demonstrate the feasibility of our aptasensing design, the OTA specific aptamer was selected as a model. The developed LC aptasensor possesses a wide linear range from 0.01 aM to 100 pM, ultra-low limit of detection (LOD) of 0.0078 aM, and quantitative recoveries of 91–103.51% for OTA in rice and grape juice samples. This study proposes a novel and universal LC-based platform for facile, ultra-sensitive, and precision sensing of hazardous analytes in real samples.

An electrochemical sensing method based on an oligonucleotide structure for ultrasensitive detection of malachite green

Herein, a simple electrochemical aptasensor with the implementation of an oligonucleotide structure is introduced for determination of malachite green (MG) as a fungicide. The oligonucleotide structure is composed of MG aptamer (Apt) and its two complementary strands (CSs). Benefiting from physical barrier and high negatively charged of the oligonucleotide structure, the proposed sensing strategy indicated improved selectivity and sensitivity for MG detection with a detection limit of 0.15 nM. Addition of MG leads to conformational change of Apt sequence, its release from the electrode surface and dismantlement of the oligonucleotide structure. So, [Fe(CN)6]3−/4− as redox marker has high accessibility to the gold electrode surface and the peak current remarkably increases. Also, the aptasensor exhibited great potential for MG detection in tap water. The designed sensing method opens a simple way for ultrasensitive detection of MG in a relatively short time.

Architecting of a biodevice based on a screen-printed carbon electrode modified with the NiONP nanolayer and aptamer in BCM-7 detection

In this study, an ultrasensitive and robust biodevice implemented on a screen-printed carbon electrode (SPCE) surface is introduced. The β-casomorphin-7 (BCM-7) peptide as an early warning sign of autism disorder is distinguished in this system. The SPCE surface was directly electrodeposited with a nanolayer of the nickel oxide nanoparticles (NiONP). In next step, an Apt sequence as a capture of the BCM-7 was strongly attached on this surface. The embedded sensing interface offered some admirable characterizes to detect the BCM-7. The obtained DPV signals were reversely proportional to the concentrations of the BCM-7 through a stable binding reaction in two working linear ranges from 0.5 × 10−9-1.5 μmol l-. Also, an unrivaled limit of detection (LOD) value of 166.6 aM was achieved that is so superior by other reported methods in the BCM-7 sensing. This hand-held biodevice was satisfactorily tested for the BCM-7 detection in human urine and blood sample with an average recovery rate of ∼101.87 %. More importantly, this strategy is free from labeling steps, complex sample processing and interference from common biomolecules in blood or urine. Due to the inherent advantages of the SPCE and the NiONP, utilizing this facile sensing interface may be an ideal choice in constructing of the ultrasensitive biodevice with low cost for distinguishing of the autism disorder.

AgNPs/QDs@GQDs nanocomposites developed as an ultrasensitive impedimetric aptasensor for ractopamine detection.

Developing easy-to-use and miniaturized sensors for in-field monitoring of targets which is related to human health is necessary. Ractopamine (RAC) is a feed additive with serious side effects that is forbidden in many countries. This study reports the fabrication of an impedimetric aptasensor for ultrasensitive and selective detection of the RAC in human biological fluids. Accordingly, an efficient nanocomposites was synthesized by a beneficial combination of graphene quantum dots (GQDs), quantum dots (QDs) and silver nanoparticles (AgNPs) for modifying a glassy carbon electrode (GCE). This nanocomposite is promising to present a synergistic effect in the increase of the active surface area of the modified electrode to more load the biocapture of the target. Next, the RAC-binding aptamer (Apt) was attached to the AgNPs/QDs@GQDs/GCE surface and a sensitive layer for the RAC detection was embedded. A RAC-Apt complex was formed upon adding the RAC and the changes of the electrochemical behavior were studied by some electrochemical techniques such as electrochemical impedance spectroscopy (EIS). Under optimal conditions, the charge transfer resistance (Rct) value was increased linearly with increasing of the RAC concentrations in the range of 1 fM to 901.4nM. Limit of detection (LOD) was calculated to be 330 aM which is superior by other reported electrochemical methods in the RAC sensing. The applicability of the aptasensor was tested in human urine and blood serum as the real samples and satisfactory results of specificity were achieved. It seems that the proposed strategy not only provides a new ultrasensitive strategy for RAC detection but also expands the application of the sensing interface to develop other aptasensors by changing the Apt sequence.

Designing of an ultrasensitive BCM-7 aptasensor based on an SPCE modified with AuNR for promising distinguishing of autism disorder

Abstractly, in this study, an aptasensor is introduced based on a platform consisting of the gold nanorod (AuNR) on a screen printed carbon electrode (SPCE) surface. The aptasensor is applied for detection of the β-casomorphin (BCM-7) as a promising biomarker of autism disorder. The NH2-Apt sequence is directly immobilized onto the AuNR/SPCE surface by formation of a chemisorption bond between the amine-Au groups. By incubation of the BCM-7 onto the aptasensor surface, the aptasensor directed against BCM-7 and cleverly formed a target/Apt complex to produce a measurable electrical current change. The aptasensor shows linearity over the range of 1 fmol L−1 to 25 nmol L−1 with a limit of detection (LOD) of 334 amol L−1. Furthermore, the function of the aptasensor in real samples such as human urine and plasma samples is evaluated. The achieved satisfactory results are mainly due to three main reasons including (1) the large specific surface area of the AuNR which forms a 3D network on the SPCE surface to capture more Apt sequences at the sensing interface, (2) utilizing Apt as the BCM-7 receptor with inherent unique properties to produce a synergetic effect with the AuNR, and finally, (3) effective using screen printing technology with the fantastic capability to less cost of the aptasensor preparation. There is hope that miniaturization of the proposed aptasensor may aid future efforts to detect autism symptoms as early as infancy under clinical conditions in real-world.

Recognition and description of synergy conditions in team work in view of the grounded theory

Developing an organisation of the future requires growing potential that will create the foundations for entrepreneurship, innovative activities and sustainable development. Every unit in the organisation's structure is to be a part of this but only team work gives the opportunity to engage the multi-directional potentials in the process. The enhanced cooperation enables any team to achieve exceptional results and the synergy effect. In this context, the value for the development of an organisation is the ability to recognise this phenomenon and indicate the circumstances of its occurrence. In the article, the grounded theory strategy applied in the research, is presented as a method enabling recognition and description of synergy conditions in a team. And based on the methodology, the author aims at explaining the factors deciding on the quality of team work. Given, however, the multidimensional nature of the research on synergy, the main objective of the article is to outline the developed research procedure first stage’s endeavours and to discuss the qualitative findings. The employed approach allow keeping apt sequence of delivering empirical evidences proving that synergy is a function of state. The applied concept of grounded theory enabled the researcher to see the key definitions and to indicate variables that would explain the conditions influencing the effectiveness of teams functioning. The ability to present the first part of the findings is, in fact, an opportunity to fortify the voiced arguments that the creative approach, entrepreneurial activity, and social capital are vital to establish the synergy effect in team work. Furthermore, the findings clearly support the concept that synergy is a feature of a team that emerges at specific time, when the team reaches a particular level of relation quality. On top of that, the acquired findings also present that team proficiency and potential growth are, at all times, strengthened by the ability to perceive and implement the synergy effect.

Preliminary application of amide proton transfer imaging signal in acute ischemic stroke

To apply amide proton transfer magnetic resonance imaging (APT-MRI) technique to acute ischemic stroke, and to discuss clinical values of the APT-MRI main parameter - MTRasym(3.5 ppm). A total of 18 emergency admitted patients, with acute ischemic stroke from Beijing Hospital, were recruited from March to November 2015. Eventually, 18 patients (male 14, female 4, average age 72 years old, age ranged from 52 to 92 years, symptom onset ≤24 hours, 2 patients≤ 6 hours) were included in this study. All patients underwent MR scanning on the brain at 3.0 Tesla. Besides the conventional scans for stroke, the APT sequence (saturation time 0.8 s, saturation power 2 μT) was performed. APT weighted (APTW) images were calculated using magnetization transfer ratio asymmetry at 3.5 ppm with respect to water. MRI signals in APTW, diffusion weighted images (DWI) and fluid attenuated iversion recovery (FLAIR) were visually evaluated. Shapiro-Wilk tests were performed to find out the distribution of MTRasym(3.5 ppm) in the whole sample, infarction regions, and normal appearing white matter (NAWMs), respectively. Detrended normal P-P plots were also used to evaluate its normality. Paired-t test and two-sample t test were applied to compare the mean MTRasym(3.5 ppm) of infarction regions with that of contra lateral NAWMs. (1) High signal intensity was observed in DWI and FLAIR in all 18 patients. However, signal intensity was somewhat different in APTW. 13 patients were found decreased APT effect, and 5 patients were with equal APT effect in infarction regions. In NAWMs, 14 patients were found equal APT effect, and 4 patients were with decreased APT effect. (2) MTRasym(3.5 ppm)s were proven, with Shapiro-Wilk test, to distribute normally within the whole sample (W=0.964, P=0.288), infarction regions (W=0.962, P=0.645), and NAWMs(W=0.929, P=0.187). However, its normality was not so satisfactory due to greater deviations (absolute value>0.06) of some cases in detrended normal P-P plots. (3) Paired-t test analysis showed that MTRasym(3.5 ppm) in infarction regions (-0.035%±1.020%) was significantly lower than that in NAWMs (0.386%±0.790%) (t=-2.273, P=0.036). However, there was no statistically significant difference between infarction regions and NAWMs when two sample t-test was performed (t=-1.386, P=0.175). MTRasym(3.5 ppm) can be used to evaluate acute ischemic stroke with a broad future scope in clinical.

Preparation of Multivalent Carbohydrate Mimetics Based on Enantiopure 1,2‐Oxazines by Sonogashira Coupling and Subsequent Reductive Ring‐Opening

AbstractA modular approach to aryl C‐glycosides and their multivalent analogues is presented. Sonogashira coupling reactions connected the key compounds, enantiopure bicyclic 1,2‐oxazine derivatives bearing p‐bromophenyl substituents, with alkynes. Subsequent hydrogenolyses or a combination of hydrogenolysis and samarium diiodide mediated reductions converted the coupling products into new unnatural amino carbohydrate mimetics with a D‐talose configuration. By Glaser coupling reactions we obtained products with a 1,3‐diyne linker and divalent compounds derived therefrom. Through Sonogashira reactions of compound 8 with several iodobenzene derivatives, di‐, tri‐, and tetravalent compounds were prepared in high yields. The subsequent reductive processes converted the 1,2‐oxazine moieties into the corresponding aminopyrans. Severe purification problems were solved in most cases by following the appropriate strategies as an apt sequence of steps or by acetylation of the intermediates.

Psoralen photo-cross-linking by triplex-forming oligonucleotides at multiple sites in the human rhodopsin gene.

Targeting DNA damage by triplex-forming oligonucleotides (TFOs) represents a way of modifying gene expression and structure and a possible approach to gene therapy. We have determined that this approach can deliver damage with great specificity to sites in the human gene for the G-protein-linked receptor rhodopsin, mutations of which can lead to the genetic disorder autosomal dominant retinitis pigmentosa. We have introduced DNA monoadducts and interstrand cross-links at multiple target sites within the gene using TFOs with a photoactivatable psoralen group at the 5'-end. The extent of formation of photoadducts (i.e., monoadducts and cross-links) was measured at target sites with a 5'-ApT sequence at the triplex-duplex junction and at a target site with 5'-ApT and 5'-TpA sequences located four and seven nucleotides away, respectively. To improve psoralen reactivity at more distant sites, psoralen moieties were attached to TFOs with nucleotide "linkers" from two to nine nucleotides in length. High-affinity binding was maintained with linkers of up to 10 nucleotides, but affinities tended to decrease somewhat with increasing linker length due to faster dissociation kinetics. DNase I footprinting indicated little, if any, interaction between linkers and the duplex. Psoralen-TFO conjugates formed DNA cross-links with high efficiency (56-65%) at 5'-ApT sequences located at triplex junctions. At a 5'-ApT site four nucleotides away, the efficiency varied with linker length; a four-nucleotide linker gave the highest efficiency. Duplexes with 5'-TpA and 5'-ApT sites two nucleotides away, in otherwise identical sequences, were cross-linked with efficiencies of 56 and 38%, respectively. These results indicate that TFO-linker-psoralen conjugates allow simultaneous, efficient targeting of multiple sites in the human rhodopsin gene.

Sequence specificity of DNA-psoralen photoproduct formation in supercoiled plasmid DNA (pUC19)

Supercoiled pUC19 DNA, photoreacted with psoralen derivatives (xanthotoxin (8-methoxypsoralen, 8-MOP), 4,5′,8-trimethylpsoralen (TMP) and angelicin), influences the enzymatic activity of restriction enzymes in a different manner, although all the enzymes employed contain, within their recognition sites, suitable nucleic acid bases for photoproduct formation. The activity of the enzymes is strongly influenced by the photomodification of thymine residues within their recognition sites. 5′-TpA sequences favour intercalation as an essential prerequisite of the photoreaction, while 5′ApT sequences do not. This, in turn, influences photoproduct formation and the inhibition of the action of the restriction enzymes KpnI, SspI, DraI and RsaI, but not EcoRI and BamHI. The inhibitory effect is independent of the number of cleavage sites and also of whether monoaddition products or crosslinks are formed. Psoralen intercalation alone does not affect the activity of the restriction enzymes used.

Actinomycin D facilitates transition of AT domains in molecules of sequence (AT)nAGCT(AT)n to a DNAse I detectable alternating structure.

The interaction of actinomycin D with (AT)nAGCT(AT)n (where n = 2, 3, or 4) was investigated using a combination of imino proton NMR and DNAse I digestion. The stoichiometry of the interaction appears to be one:one with the actinomycin chromophore intercalated between the two GC base pairs. This binding event facilitates the conversion of the flanking repetitive AT regions to an alternating conformation characterized by induced sensitivity of the ApT sequences to attack by DNAse I. The neighboring TpA sequences do not exhibit rate changes as a function of binding of the drug. The potential relevance of such ligand induced DNA structural alterations is discussed.