Nano Fluorescent Probes Used In Research And Application Of Biologically Active Molecules

The new type of fluorescent nanoprobe with its special quantum size effects and small dimension effects exhibits many different optical characteristics compared to the homogeneous single-molecule or large object, and thus has attracted great attention in the applied research field on analytical chemistry. At present, the widely investigated nanomaterial is the semiconductor nanoparticles which are also called quantum dots (QDs). When their size is comparable to the size of Bohr diameter for exciton, they exhibit special physical and chemical properties which possess the quantum size effects, dielectric confinement effects, surface effects, macroscopic quantum tunneling effect and so on. So the quantum dots with these special optical characteristics applied in the fluorescent biological analytical applications have become a wide research focus.Compared to conventional organic dyes, QDs possess many advantages, such as narrower emission spectra, tunable maximum emission wavelength with changeable sizes and compositions, photostability, high brightness, long fluorescence lifetime and biocompatibility, which have been successfully used as fluorescent probe or sensor in the imaging of biological samples and cells. For many decades, scientists have recognized the power of incorporating biological principles and molecules into the design of artificial devices. Following recent advances in the growing field of nanotechnology, nanomaterials can be designed as exquisitely sensitive chemical and biological sensors. Nanosensors with immobilized bioreceptor probes that are selective for target analyte molecules are called nanobiosensors which generally consist of a biosensitive part that can either contain biological recognition elements or be made of biological recognition elements covalently attached to the transducer. The interaction between the target analyte and bioreceptor is designed to produce a physicochemical perturbation on nanobiosensors that can be converted into a measurable effect such as an optical or electrical signal. And thus the effects of nanomaterials on biological systems as well as the function research of biomolecules can be realized. Recent advances in the application of nanomaterials as well as their complexes with biomolecules in biosensing and biodetection has attracted great attention. For example, QDs with its good optical characteristics, high catalytic effects and high electron transfer efficiency has been used as a wonderful material in enzyme-based biological analyses and applications. Therefore, the design of elegant new assembled nanobiosensors for realizing the analysis and determination of bioactive molecules in vivo or in vitro has become a great challenge to the scientific workers.Based on the merits of water-soluble CdTe QDs with good optical characteristics and biocompatibility, we newly assembled two fluorescent nanobiosensors for the direct determination of glucose in biological systems.First, a new nanobiosensor QDs-ConA-β-CDs-AuNPs was assembled for the direct determination of glucose in human serum based on the fluorescence resonance energy transfer (FRET) between CdTe QDs and gold nanoparticles (AuNPs). The sensing mechanism is based on the switching off FRET through the high specific recognition of concanavalin A (ConA) to glucose. In the presence of glucose, the AuNPs-β-CDs segment of the nanobiosensor is displaced by glucose which competes withβ-CDs on the binding sites of ConA, resulting in the fluorescence recovery of the quenched QDs. The results show that the linear range of this method is 0.10μM~50μM with the detection limit as low as 50 nM, and has excellent selectivity for glucose over other sugars and most biological species present in serum. The nanobiosensor was applied directly to determine glucose present in normal adult human serum, and the recovery and precision of the method were satisfactory.Second, a new nanocomplex CdTe QDs-GOx was assembled, and thus realized the great enhanced enzymatic activity, widened the active temperature range of glucose oxidase (GOx) and can be used as a nanosenor for the simultaneous determination of glucose. The determination mechanism is put forward based on the electron transfer mechanism on the fluorescence quenching of CdTe QDs. The results indicate that the lower value of the Michaelis-Menton constant is estimated to be 0.45 mM/L, which shows the greatest enhanced enzymatic activity measured by far. Besides, the GOx enzyme conjugated on the CdTe QDs obtains better thermal stability at 20-80°C and keeps the maximum activity in the wide range of 40-50°C. The linear range for glucose sensing of this method is 5.0μM~1.0 mM with the detection limit of 0.10μM, and would provide a new approach for the direct assay on the lower level of glucose in complicated biological systems.