Blank peak current-suppressed electrochemical aptameric sensing platform for highly sensitive signal-on detection of small molecule

In this contribution, an electrochemical aptameric sensing scheme for the sensitive detection of small molecules is proposed using adenosine as a target model. A ferrocene (Fc)-functionalized thiolated aptamer probe is adapted and immobilized onto an electrode surface. Introducing a recognition site for EcoRI into the aptamer sequence not only suppresses the peak current corresponding to blank sample but also provides a signal-on response mechanism. In the absence of adenosine, the aptamer can fold into a hairpin structure and form a cleavable double-stranded region. Fc is capable of being removed from electrode surface by treatment with endonuclease, and almost no peak current is observed. The adenosine/aptamer binding induces the conformational transition of designed aptamer, dissociating the cleavable double-stranded segment. Therefore, the integrated aptamer sequence is maintained when exposing to endonuclease, generating a peak current of Fc. Utilizing the present sensing scheme, adenosine even at a low concentration can give a detectable current signal. Thus, a detection limit of 10−10 M and a linear response range from 3.74 × 10−9 to 3.74 × 10−5 M are achieved. The proposed proof-of-principle of a novel electrochemical sensing is expected to extend to establish various aptameric platforms for the analysis of a broad range of target molecules of interest

Folding of the cocaine aptamer studied by EPR and fluorescence spectroscopies using the bifunctional spectroscopic probe Ç

The cocaine aptamer is a DNA molecule that binds cocaine at the junction of three helices. The bifunctional spectroscopic probe Ç was incorporated independently into three different positions of the aptamer and changes in structure and dynamics upon addition of the cocaine ligand were studied. Nucleoside Ç contains a rigid nitroxide spin label and can be studied directly by electron paramagnetic resonance (EPR) spectroscopy and fluorescence spectroscopy after reduction of the nitroxide to yield the fluoroside Çf. Both the EPR and the fluorescence data for aptamer 2 indicate that helix III is formed before cocaine binding. Upon addition of cocaine, increased fluorescence of a fully base-paired Çf, placed at the three-way junction in helix III, was observed and is consistent with a helical tilt from a coaxial stack of helices II and III. EPR and fluorescence data clearly show that helix I is formed upon addition of cocaine, concomitant with the formation of the Y-shaped three-way helical junction. The EPR data indicate that nucleotides in helix I are more mobile than nucleotides in regular duplex regions and may reflect increased dynamics due to the short length of helix I

Following Glucose Oxidase Activity by Chemiluminescence and Chemiluminescence Resonance Energy Transfer (CRET) Processes Involving Enzyme-DNAzyme Conjugates

A hybrid consisting of glucose oxidase-functionalized with hemin/G-quadruplex units is used for the chemiluminescence detection of glucose. The glucose oxidase-mediated oxidation of glucose yields gluconic acid and H2O2. The latter in the presence of luminol acts as substrate for the hemin/G-quadruplex-catalyzed generation of chemiluminescence. The glucose oxidase/hemin G-quadruplex hybrid was immobilized on CdSe/ZnS quantum dots (QDs). The light generated by the hybrid, in the presence of glucose, activated a chemiluminescence resonance energy transfer process to the QDs, resulting in the luminescence of the QDs. The intensities of the luminescence of the QDs at different concentrations of glucose provided an optical means to detect glucose

Sensitive fluorescence detection of nucleic acids based on isothermal circular strand-displacement polymerization reaction

Here we have developed a sensitive DNA amplified detection method based on isothermal strand-displacement polymerization reaction. This method takes advantage of both the hybridization property of DNA and the strand-displacement property of polymerase. Importantly, we demonstrate that our method produces a circular polymerization reaction activated by the target, which essentially allows it to self-detect. Functionally, this DNA system consists of a hairpin fluorescence probe, a short primer and polymerase. Upon recognition and hybridization with the target ssDNA, the stem of the hairpin probe is opened, after which the opened probe anneals with the primer and triggers the polymerization reaction. During this process of the polymerization reaction, a complementary DNA is synthesized and the hybridized target is displaced. Finally, the displaced target recognizes and hybridizes with another probe, triggering the next round of polymerization reaction, reaching a target detection limit of 6.4 × 10−15 M

Chromogenic and Fluorogenic Probes for the Detection of Illicit Drugs

[EN] The consumption of illicit drugs has increased exponentially in recent years and has become a problem that worries both governments and international institutions. The rapid emergence of new compounds, their easy access, the low levels at which these substances are able to produce an effect, and their short time of permanence in the organism make it necessary to develop highly rapid, easy, sensitive, and selective methods for their detection. Currently, the most widely used methods for drug detection are based on techniques that require large measurement times, the use of sophisticated equipment, and qualified personnel. Chromo- and fluorogenic methods are an alternative to those classical procedures.We thank the Spanish Government [projects MAT2015-64139-C4-1-R and AGL2015-70235-C2-2-R (MINECO/FEDER)] and the Generalitat Valenciana (project PROMETEOII/2014/047) for support. S.E.S thanks the Ministerio de Economia y Competitividad for his Juan de la Cierva contract. 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Supramolecular Sensors for Opiates and Their Metabolites. Journal of the American Chemical Society, 139(42), 14954-14960. doi:10.1021/jacs.7b0637

Aptamer-Functionalized Nano-Biosensors

Nanomaterials have become one of the most interesting sensing materials because of their unique size- and shape-dependent optical properties, high surface energy and surface-to-volume ratio, and tunable surface properties. Aptamers are oligonucleotides that can bind their target ligands with high affinity. The use of nanomaterials that are bioconjugated with aptamers for selective and sensitive detection of analytes such as small molecules, metal ions, proteins, and cells has been demonstrated. This review focuses on recent progress in the development of biosensors by integrating functional aptamers with different types of nanomaterials, including quantum dots, magnetic nanoparticles (NPs), metallic NPs, and carbon nanotubes. Colorimetry, fluorescence, electrochemistry, surface plasmon resonance, surface-enhanced Raman scattering, and magnetic resonance imaging are common detection modes for a broad range of analytes with high sensitivity and selectivity when using aptamer bioconjugated nanomaterials (Apt-NMs). We highlight the important roles that the size and concentration of nanomaterials, the secondary structure and density of aptamers, and the multivalent interactions play in determining the specificity and sensitivity of the nanosensors towards analytes. Advantages and disadvantages of the Apt-NMs for bioapplications are focused