First report of the use of a saxitoxin-protein conjugate to develop a DNA aptamer to a small molecule toxin

Saxitoxin (STX) is a low molecular weight neurotoxin mainly produced by certain marine dinoflagellates that, along with its family of similarly related paralytic shellfish toxins, may cause the potentially fatal intoxication known as paralytic shellfish poisoning. Illness and fatality rates are low due to the effective monitoring programs that determine when toxins exceed the established regulatory action level and effectuate shellfish harvesting closures accordingly. Such monitoring programs rely on the ability to rapidly screen large volumes of samples. Many of the screening assays currently available employ antibodies or live animals. This research focused on developing an analytical recognition element that would eliminate the challenges associated with the limited availability of antibodies and the use of animals. Here we report the discovery of a DNA aptamer that targets STX. Concentration-dependent and selective binding of the aptamer to STX was determined using a surface plasmon resonance sensor. Not only does this work represent the first reported aptamer to STX, but also the first aptamer to any marine biotoxin. A novel strategy of using a toxin-protein conjugate for DNA aptamer selection was successfully implemented to overcome the challenges associated with aptamer selection to small molecules. Taking advantage of such an approach could lead to increased diversity and accessibility of aptamers to low molecular weight toxins, which could then be incorporated as analytical recognition elements in diagnostic assays for foodborne toxin detection. The selected STX aptamer sequence is provided here, making it available to any investigator for use in assay development for the detection of STX.

Shape corrections to exchange-correlation potentials by gradient-regulated seamless connection of model potentials for inner and outer region

Shape corrections to the standard approximate Kohn-Sham exchange-correlation (xc) potentials are considered with the aim to improve the excitation energies (especially for higher excitations) calculated with time-dependent density functional perturbation theory. A scheme of gradient-regulated connection (GRAC) of inner to outer parts of a model potential is developed. Asymptotic corrections based either on the potential of Fermi and Amaldi or van Leeuwen and Baerends (LB) are seamlessly connected to the (shifted) xc potential of Becke and Perdew (BP) with the GRAC procedure, and are employed to calculate the vertical excitation energies of the prototype molecules N-2, CO, CH2O, C2H4, C5NH5, C6H6, Li-2, Na-2, K-2. The results are compared with those of the alternative interpolation scheme of Tozer and Handy as well as with the results of the potential obtained with the statistical averaging of (model) orbital potentials. Various asymptotically corrected potentials produce high quality excitation energies, which in quite a few cases approach the benchmark accuracy of 0.1 eV for the electronic spectra. Based on these results, the potential BP-GRAC-LB is proposed for molecular response calculations, which is a smooth potential and a genuine "local" density functional with an analytical representation. (C) 2001 American Institute of Physics.