KEYWORDS: Nanoparticles, Nanomaterials, Gold, Energy transfer, Luminescence, Metals, Fluorescence resonance energy transfer, Magnesium, Analytical research, Molecular energy transfer
Energy transfer from organic fluorophores to small metal nanoparticles is being used as a molecular beacon tool to monitor the kinetic processes of the hammerhead ribozyme. This marks the first time that nanomaterials have been used to monitor ribozyme kinetics. The quantum efficiency of energy transfer from the fluorophore to the gold nanoparticle follows a distance dependence behavior, which allows the real-time characterization of ribozyme complex structure and cleavage kinetics. The rate of cleavage for our ribozyme at pH=6.5 and 37°C is measured to be on the order of 10-2 min-1, which is the correct order of magnitude for similar ribozymes at this pH in the literature.
Multi-pathogen biosensors that take advantage of sandwich immunoassay detection schemes and utilize conventional fluorescent dye reporter molecules are difficult to make into extremely compact and autonomous packages. The development of a multi-pathogen, immunoassay-based, fiber optic detector that utilizes varying sized fluorescent semiconductor quantum dots (QDs) as the reporter labels has the potential to overcome these problems. In order to develop such a quantum dot-based biosensor, it is essential to demonstrate that QDs can be attached to antibody proteins, such that the specificity of the antibody is maintained. We have been involved in efforts to develop a reproducible method for attaching QDs to antibodies for use in biodetection applications. We have synthesized CdSe/ZnS core-shell QDs of differing size, functionalized their surfaces with several types of organic groups for water solubility, and covalently attached these functionalized QDs to rabbit anti-ovalbumin antibody protein. We also demonstrated that these labeled antibodies exhibit selective binding to ovalbumin antigen. We characterized the QDs at each step in the overall synthesis by UV-VIS absorption spectroscopy and by picosecond (psec) transient photoluminescence (TPL) spectroscopy. TPL spectroscopy measurements indicate that QD lifetime depends on the size of the QD, the intensity of the optical excitation source, and whether or not they are functionalized and conjugated to antibodies. We describe details of these experiments and discuss the impact of our results on our biosensor development program.
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