Quantum Dots - Composite Probe Of Biomolecules In Bioanalysis Applications

Semiconductor quantum dots (QDs) are nanoparticles that have attracted widespread interest in biology and medicine due to their unique optical and electronic properties. These nanoparticles possess high PLQY and photostability, as well as size-dependent and tunable photoluminescence with narrow emission bandwidth and broad excitation, which make them highly promising fluorescent labels for both in vitro and in vivo studies. Most of these applications are based on specific interaction between biomolecules. So construction of QD-biconjugates, to endue QDs with specific targeting effect, high fluorescence intensity and stability, low non-specific absorption, is critical to bio-application of QDs.With the development of medical technology, medical detection requested inspection instruments have higher sensitivity, shorter detection time, and smaller volume. The microfluidic chip technology has the advantages that could satisfy these requirements. This miniaturized analysis technology is particularly useful in many biomedical fields, e.g. gene sequeching, protein pattern analysis. With microfluidic technology constantly mature and production cost continuous reduced, this new technology will enter large-scale testing field of conventional biomedicine, replace the cumbersome inspection equipments and complicated operations. However, microfluidic biochip technology is still faced with several challenges, such as relatively low sensitivity and specificity.High-quality QDs were successfully prepared directly in aqueous phase assisted by microwave irradiation in our laboratory. Based on this advantage, we attempted to combine QDs with microfluidic biochip technology. The excellent fluorescent properties of QDs will increase the sensitivity of microfluidic biochips. In this paper, a serial of QD-based bio-nanoprobes were constructed, and applied in the microfluidic biochip. The details are as follows:1. A novel microfluidic biochip was constructed.This microfluidic biochip was consisted of PDMS and substrate-slide. This biochip allowed facile patterning of biomolecules and subsequent biomolecular recognition. It was proved that this new method had many advantages, such as low-cost, easy to operate and could analyze different kinds of targets in varied concentrations. Compared with microarray technology, micofluidic biochip does not require expensive automated spotting robots as well as strict reaction conditions such as humidity and temperature control.2. QD-avidin bioconjugate was prepared and applied in the microfluidic protein chip.QD-avidin bioconjugate was prepared based on electrostatic attraction. This fluorescent nanoprobe was applied in the microfluidic protein chip, and cancer markers were assayed with high sensitivity and selectivity. Experimental conditions were optimized to reduce the non-specific binding and enhance the signal-to-noise ratio. It bore out that, compared with organic dyes, QDs exhibited higher signal intensity, better stability and higher sensitivity in microfluidic protein chip. We thus expect this detection system will be a promising tool for molecular diagnostics and proteomics.3. QD-IgG bioconjugate was prepared and applied in the microfluidic protein chip.We developed a microfluidic protein chip for an ultrasensitive and multiplexed assay of cancer biomarkers. QDs were employed as fluorescent signal amplifiers to improve the detection sensitivity. Secondary antibodies (goat antimouse IgG) were conjugated to luminescent QDs to realize a versatile fluorescent probe that could be used for multiplexed detection in both sandwich and reverse phase immunoassays. We found that our microfluidic protein chip not only possessed ultrahigh femtomolar sensitivity for cancer biomarkers, but was selective enough to be directly used in serum. This protein chip thus combines the high-throughput capabilities of a microfluidic network with the high sensitivity and multicolor imaging ability offered by highly fluorescent QDs, which can become a promising diagnostic tool in clinical applications.4. QD-DNA-biotin-strepavidin bioconjugate was prepared and applied in the microfluidic protein chip.We have demonstrated a novel, DNA-bridged, strategy for assemble proteins at the surface of fluorescent semiconductor QDs. As a proof-of-concept, we designed and synthesized a QD-DNA-biotin-STV bioconjugate that can serve as a generic fluorescent nanoprobe for ultrasensitive protein detection. We have demonstrated the preparation of this QD-DNA-biotin-STV conjugate is convenient and stable, with minimal perturbation to the optical properties of QDs. We then employ this bioconjugate as a fluorescent nanoprobe for ultrasensitive detection of cancer biomarkers with a microfluidic protein chip.5. QD-DNA bioconjugate was prepared and applied in microfluidic DNA chip.QD-DNA bioconjugate was prepared by ligand exchange. Fluorescence resonance energy transfer (FRET) system based on two-color QDs was constructed for target DNA detection. Besides, this QD-DNA bioconjugate served as a fluorescent nanoprobe for DNA detection with high sensitivity. What's more important, this DNA detection system was able to discriminate the complementary DNA, single base-mismatch DNA, and non-complementary DNA.6. A label-free glucose detection system based on QDs was designed.We demonstrated that the photoluminescence (PL) of CdTe/CdS QDs was sensitive to hydrogen peroxide (H2O2). With D-glucose as the substrate, H2O2 that intensively quenched the QDs PL can be produced via the catalysis of glucose oxidase (GOX). In addition, the QD-based label-free glucose sensing platform was adapted to 96-well plates for fluorescent assays. The comparison with commercialized glucose meter indicated that this proposed glucose assay system is not only simple, sensitive, but also reliable and suitable for practical application.