Waterloo Institute for Nanotechnology

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Browsing Waterloo Institute for Nanotechnology by Subject "Adenosine"

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    Flow Cytometry-Assisted Detection of Adenosine in Serum with an Immobilized Aptamer Sensor
    (American Chemical Society, 2010-05-15) Huang, Po-Jung Jimmy; Liu, Juewen
    Aptamers are single-stranded nucleic acids that can selectively bind to essentially any molecule of choice. Because of their high stability, low cost, ease of modification, and availability through selection, aptamers hold great promise in addressing key challenges in bioanalytical chemistry. In the past 15 years, many highly sensitive fluorescent aptamer sensors have been reported. However, few such sensors showed high performance in serum samples. Further challenges related to practical applications include detection in a very small sample volume and a low dependence of sensor performance on ionic strength. We report the immobilization of an aptamer sensor on a magnetic microparticle and the use of flow cytometry for detection. Flow cytometry allows the detection of individual particles in a capillary and can effectively reduce the light scattering effect of serum. Since DNA immobilization generated a highly negatively charged surface and caused an enrichment of counterions, the sensor performance showed a lower salt dependence. The detection limits for adenosine are determined to be 178 μM in buffer and 167 μM in 30% serum. Finally, we demonstrated that the detection can be carried out in 10 μL of 90% human blood serum.
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    Molecularly Imprinted Polymers with DNA Aptamer Fragments as Macromonomers
    (American Chemical Society, 2016-03-16) Zhang, Zijie; Liu, Juewen
    Molecularly imprinted polymers (MIPs) are produced in the presence of a template molecule. After removing the template, the cavity can selectively rebind the template. MIPs are attractive functional materials with a low cost and high stability, but traditional MIPs often suffer from low binding affinity. This study employs DNA aptamer fragments as macromonomers to improve MIPs. The DNA aptamer for adenosine was first split into two halves, fluorescently labeled, and copolymerized into MIPs. With a fluorescence quenching assay, the importance of imprinting was confirmed. Further studies were carried out using isothermal titration calorimetry (ITC). Compared to the mixture of the free aptamer fragments, their MIPs doubled the binding affinity. Each free aptamer fragment alone cannot bind adenosine, whereas MIPs containing each fragment are effective binders. We further shortened one of the aptamer fragments, and the DNA length was pushed to as short as six nucleotides, yielding MIPs with a dissociation constant of 27 mu M adenosine. This study provides a new method for preparing functional MIP materials by combining high-affinity biopolymer fragments with low-cost synthetic monomers, allowing higher binding affinity and providing a method for signaling binding based on DNA chemistry.