Preparation And Application Of Carbon Nanotubes-based Affinity Adsorbent

Recently, carbon nanotubes (CNTs) exhibit good application potentials in biological fields.They have been extensively studied in the areas of biosensors, drug-delivery systems and immobilization and purification of biological macromolecules. However, problems exist with the practical use of CNTs. First of all, strong hydrophobic property of CNTs not only causes non-specific adsorption of proteins, but also leads to the the aggregation of CNTs, thus significantly reducing the effective surface area. Besides, CNTs are very lightweight, centrifugation with long time and high speed is usually used to collect CNTs, which is time and energy consuming. This paper attempted to modify CNTs to solve the problems above. Subseqently, affinity ligand was immobilized on modified CNTs to study the affinity purification capacity of this adsorbent for target protein. The work in this thesis includes:(1) CNTs were acid treated, then modified with magnetic nanoparticles, coated with polyethyleneimine (PEI) and lastly functionalized with protein A to prepare PEI modified carbon nanotubes-based affinity adsorbent (MMWCNTs-PEI-PA). The stability of PEI modification layer was studied. The modified CNTs were characterized by SEM, FTIR and VSM. The results showed that PEI modification layer exhibited good stability, even after being treated with a high concentration of salt. MMWCNTs-PEI-PA showed good dispersion. Magnetic analysis indicated that the adsorbent was superparamagnetic and could be collected conveniently and rapidly in the presence of an external magnetic field.(2) The performance for immunoglobulin G (IgG) adsorption and purification of MMWCNTs-PEI-PA was studied. PEI framework on the adsorbent caused non-specific adsorption for acidic proteins via electrostatic attraction. However, such non-specific adsorption was eliminated successfully by increasing the salt concentration of the adsorption solution. Next, adsorption kinetics and isotherm were studied. MMWCNTs-PEI-PA showed rapid adsorption rate and high affinity for IgG adsorption. The time for reaching adsorption equilibrium was about40min. The association constant of MMWCNTs-PEI-PA for IgG was1.6×106L/mol, and the maximum adsorption capacity was30.55mg/g. IgG adsorption experiments from human serum showed that this adsorbent presented high specificity towards IgG with a final purity of over90%under optimum conditions. In addition, the adsorption capacity of the absorbent for IgG was declined only slightly and the specificity showed little change after ten cycles. (3) MMWCNTs-Dex-PA was prepared by modifying PEI-coated CNTs with dextran and then with protein A. The performance for IgG adsorption and purification of MMWCNTs-Dex-PA was studied. Dextran modification layer significantly inhibited the non-specific adsorption caused by PEI framework. The association constant of MMWCNTs-Dex-PA for IgG was1.4×106L/mol. IgG adsorption experiments from human serum showed that MMWCNTs-Dex-PA also presented high specificity towards IgG and excellent reusability.