| With the accomplishment of the gene sequencing of Human Genomic Project (HGP), increasing attention is focused on proteomic research. Currently, one of the major bottle necks encountered in proteomic research is to deplete high abundant proteins and enrich low abundant proteins from complex biological samples. In order to resolve the problem, it is necessary to develop some novel separation methods and technologies. Herein, we combined immobilized metal affinity with magnetic separation to remove the abundant protein (bovine hemoglobin, BHb) in the bovine blood. On the other hand, we have synthesized mesoporous silica coated multiwall carbon nanotube to separate cytochrome c (Cyt c) from the mixture of bovine serum albumin (BSA), BHb and Cyt c.(1) In chapter 1, we have summarized the current progress in metal chelation technique with magnetic separation, especially the separation and purification of protein. Considering the abundant protein BHb existing in bovine blood, so it is meaningful to remove the abundant protein using the technology. On the other hand, the mesoporous materials have been widely used in the biotechnology field, the protein immobilization on the mesoporous materials has been made great progress.However, it has seldomly been used for the protein separation, and the size of pore is very important for the separation of proteins.(2) In chapter 2, we have synthesized the Fe3O4 as the matrix, and explored the factors on the morphology of Fe3O4 by controlling the amount of PVP. Furthermore, we also coat the Fe3O4 with silica, and discuss the difference between the ethanol-water and isopropal-water systems. Next, the trichloro(4-chloromethylphenyl) silane was immobilized on them and after reacted with iminodiacetic acid (IDA) and charged with Cu2+. The obtained magnetic silica microspheres with immobilized Cu2+ were applied for the absorption of BHb and the removal of BHb from bovine blood. The size, morphology and magnetic property of the resulting magnetic micro(nano) spheres were investigated by scanning microscopy (SEM), transmission electron microscopy (TEM), X-ray diffraction (XRD) and vibrating sample magnetometer (VSM).(3) In chapter 3, a novel route for the preparation of magnetite (Fe3O4) nanoparticles with immobilized metal affinity ligand iminodiacetic acid (IDA) charged with Cu2+ was developed. It is interesting that magnetic nanoparticles modified with metal ligands showed property of magnetic colloid photonic crystal. Furthermore, it could efficiently remove the abundant protein bovine hemoglobin from bovine blood. It has the potential application in removing abundant protein in proteomic analysis.(4) In chapter 4, we have synthesized Cu-IDA functionalized core-satellite composite nanoparticles that possess both the superparamagnetic properties of magnetic nanoparticles and the surface chemistry of gold nanoparticles. Due to the long organic chain attached on the gold and polydopamine surface and Cu-IDA group as the "anchor" at the end for capturing target molecules, this composite material is highly specific on the removal of the abundant protein BHb in the bovine blood.(5) In chapter 5, a facile method to coat SiO2 layer onto the CNTs with the aid of the cationic surfactant CTAB was developted. Detailed characterization studies show that mesoporous CNTs@SiO2 composites possess mesostructures, high surface area, large pore volumes, and narrow pore size distributions. The adsorption of mesoporous CNTs@SiO2 to the proteins with different molecular size like Cyt c, BSA and lysozyme (Lyz) was investigated. The adsorption results demonstrated the mesoporous CNTs@SiO2 possessing in size-selective to Cyt c and releasing easily in high salt solution. |
PDF Full Text Request