Construction And Biological Application Of Novel Nanobiological Probes

Neurological diseases and tumors are two diseases seriously impacting people’s health and life quality in the world, which have hitherto not being thoroughly overcomed. It provided opportunities for the development in diagnoses and treatment of these two diseases due to advances of nanotechnology. This thesis is focusing on the constructions and applications of new nano-bioprobes, which associates with two important issues of the diseases above: the internalization and degradation of β-amyloid peptide(Aβ) by astrocytes from CNS and the rapid sensitive detection of circulating tumor cells(CTCs).Firstly, Aβ were modified onto the surface of quantum dots(QDs) by means of physical coating, covalent coupling and biological affinity respectively. The probes were characterized through several approaches. And results showed sound maintenance of these three probes of good fluorescent properties from QDs and the immuno-biological activities of Aβ, suggesting their further applications. Then primary cultured astrocytes were applied to uptake three probes above and experimental factors were optimized. It indicated the best tracing performance of physical coating QDs-Aβ probe in cellular internalization events. Further more, this QDs-Aβ probe was used in a preliminary observation to trace the “ endocytosis- being degraded- completed degradation” process of cellular Aβ events, and consequently it proved the feasibilities of such nano-bioprobe for research purposes.Secondly, another two types of nano-bioprobes were constructed in this thesis, aiming at isolating and identifying the CTCs in a rapid, convenient and efficient way. One is named immuno-magnetic nanoparticles(IMNs) probes for cell enriching on the basis of superparamagnetic γ-Fe2O3 nanomaterials, and the other is immuno-fluorescent QDs probes that was built upon water-soluble ZnSe/ZnS QDs. Then a dye specific to Zn2+ FluoZin-3, which is able to sensitively amplify fluorescent signals was involved, providing a method of quantitative detection for CTCs. Results showed that IMNs modified by anti-EpCAM IgG could specifically capture MCF-7(a kind of breast cancer cells) in the concentrations of 1-100 cells/m L or 104 cells/mL, and the capture efficiency was above 95%. CTCs targeting QDs probe, which was prepared through mild non-copper catalyzed click chemistry reaction, could specifically identify MCF-7 cells. In addition, under optimized experimental conditions, Fluo Zin-3 could not only quantitatively analyze ZnSe/ZnS QDs through ion exchange reaction, but quantitatively detect MCF-7 cells which were enriched by IMNs and labeled with ZnSe/ZnS QDs. The linearity of this method to MCF-7 cells ranged from100 to 1000 cells/m L.In conclusion, it showed more promises that these studies could contribute to the promotion of researches on both the process and the mechanism of Aβ internalization and degradation by astrocytes, and the impetus to the development of a rapid, efficient, sensitive detection technique of CTCs. Also, it will hopefully facilitate the advances of nano-bioprobe applications in biomedical sciences.