S-Space College of Engineering/Engineering Practice School (공과대학/대학원) Dept. of Chemical and Biological Engineering (화학생물공학부) Theses (Ph.D. / Sc.D._화학생물공학부)
Biomolecule Immobilization Platforms Based on Pentafluorophenyl Acrylate Polymers
펜타플루오로페닐 아크릴레이트 고분자를 기반으로 한 생체분자 고정용 플랫폼의 제조
- 공과대학 화학생물공학부
- Issue Date
- 서울대학교 대학원
- 학위논문 (박사)-- 서울대학교 대학원 : 공과대학 화학생물공학부, 2018. 8. 차국헌.
- Functional polymers have had attention and been expected as a promising materials in a wide range of application fields such as biotechnology, photonics, and optoelectronics, and biocompatibility. The physical or chemical properties and nanostructures constituted of functional polymer should be controlled by diverse synthesis and processing methods. As more complicated platforms based on functional polymers required, reactive ester polymers were utilized for both preparation of novel polymeric precursors and for the fabrication of reactive polymer-based platforms with desired functionalities and forms. The controlled radical polymerization techniques were utilized to yield in well-defined polymers and polymer-based films and brushes. Furthermore, modification of reactive polymer platforms results in facile approaches to bio-applications by covalent immobilization of biomolecules. In this thesis, poly(pentafluorophenyl acrylate) (poly(PFPA)), one of active ester polymers, was realized in the structures of thin films and polymer brushes and considered as biomolecule immobilization platforms through post-polymerization modification methods using its high reactivity with amines. The brief introduce about reactive polymer and synthesis methods and biomolecule immobilization are in chapter 1.
In chapter 2, we investigate the mechanism in primary amine-induced post-polymerization modification of spin-cast active ester polymer thin films, comprised of poly(PFPA). The most important physical parameters in the post-modification are the molecular weight of PFPA polymers and the aliphatic chain length of primary amines. The effect of two parameters on the penetration depth as well as the exchange kinetics was systematically studied by neutron reflectivity (NR) and quartz crystal microbalance (QCM-D), accompanied by the surface morphological changes measured by an atomic force microscope (AFM) and an optical microscope (OM). The spin-cast thin films of high and low molecular weight of poly(PFPA) showed the distinctive difference originating from the primary alkyl amines of different alkyl chain length. The aliphatic chain length of primary alkyl amines dramatically influenced the penetration kinetics into low molecular weight poly(PFPA) films whereas there was no significant penetration effect on the high molecular weight films. The high molecular weight of poly(PFPA) films led to the deceleration of dissolution of amine-functionalized polymer chains in good solvent. Both alkyl chain length of primary alkyl amines and the molecular weight of poly(PFPA) affect the penetration depth and dissolution of the polymer chains from the surface of thin films, respectively.
In chapter 3, we present the synthesis of reactive polymer brushes prepared by surface-initiated (SI) RAFT polymerization of pentafluorophenyl acrylate. Dithiobenzoic acid benzyl-(4-ethyltrimethoxylsilyl) ester was used as the surface-initiated RAFT chain transfer agent (SI-CTA) and the anchoring group onto the silica particles. Poly(pentafluorophenyl acrylate) (poly(PFPA)) is known to have high and selective reactivity with amine functional groups that offers facile routes to realize diverse functions starting from the same platforms by simple post-polymerization modification with amines. Through the grafting-from approach, polymer brushes with controlled molecular weight and conformal coverage were obtained. The synthesis and utilization of reactive polymer brushes offers an easy approach in the controlled-fabrication of polymer brushes with desired functionality, which is limited by other strategies.
In chapter 4, we reported the poly(PFPA) brush-based platforms for antibody-antigen precipitation or immunoprecipitation (IP), which are routinely performed by biologists to isolate specific antigens and to identify their interactors from complex protein mixtures. The conventional approach involving agarose supports shows reasonably good antibody-binding capability due to their selective bioaffinity immobilization, but often suffers from high nonspecific binding and antibody contamination. We prepared silica particles containing poly(PFPA) brushes, prepared by the reversible addition-fragmentation chain transfer (SI-RAFT) polymerization. Upon sequential functionalization with antibodies and polyethylene glycol (PEG), it showed significantly reduced nonspecific protein adsorption and complete elimination of antibody contamination. Furthermore, by optimizing the two parameters such as molecular weight of the polymer brushes and the amount of PEG passivates, the poly(PFPA) brush-grafted particles show the highest efficiency. Taking into account their versatility and convenient features of such reactive brush platforms, the poly(PFPA) platforms have the potential to be an alternative to traditional agarose-based platforms for immunoprecipitation.
In chapter 5, we briefly introduced about poly(PFPA)-coated channel for the application of biosensors. Substrates which have amine functional groups on the surface were coated with poly(PFPA) as loop or train configuration of grafted brushes. To get more stability of poly(PFPA) films during incubation process in antibody solution, many conditions were tested and characterized by confocal images. Some functional groups in poly(PFPA) chains reacted with amine groups on substrates and the other remained groups were used for immobilization of fluorescent antibodies. Based on the fabrication of channel on the poly(PFPA)-coated substrates, we expected this simply fabricated poly(PFPA)-based platforms can be applied as biosensor for primary diagnosis.
In conclusion, reactive poly(PFPA) platforms that allow facile preparation of functional material was demonstrated from polymer films to polymer brush particles. The reactive poly(PFPA) thin films and particles based on the simple and quantitative post-modification with amine-containing molecules could be utilized for many practical applications due to the ease of control over the degree of functionalization. Our system and the strategy would provide a facile process towards functional polymer film, and polymer brushes by eliminating difficult multistep of synthesis. Furthermore, the possibility of poly(PFPA)-based platforms for the use in bioapplications such as purification and biosensing was confirmed.
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