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Human Plasma Adsorption to Biomaterials: Fundamental Level Chemical Modifications and Their Effects on Biocompatibility Open Access


Other title
Protein Adsorption
Human Plasma
Elastin-Like Polypeptides
Protein Nanoparticles
Protein Biopolymer
Type of item
Degree grantor
University of Alberta
Author or creator
Bahniuk, Markian S
Supervisor and department
Unsworth, Larry D (Chemical and Materials Engineering, Biomedical Engineering)
Examining committee member and department
Keeley, Frederick W (Department of Biochemistry & Laboratory Medicine and Pathobiology at the University of Toronto)
Uludag, Hasan (Chemical and Materials Engineering, Biomedical Engineering)
Burrell, Robert (Chemical and Materials Engineering, Biomedical Engineering)
Saldana, Marleny D. Aranda (Agricultural, Life and Environmental Sciences)
Adeeb, Samer (Civil and Environmental Engineering)
Department of Biomedical Engineering

Date accepted
Graduation date
2017-11:Fall 2017
Doctor of Philosophy
Degree level
The non-specific adsorption of proteins to biomaterials is a process that begins instantly upon implantation and results in the formation of a protein-rich layer coating the biomaterial surface. The consequences of this complex phenomenon are far-reaching and may affect both the biomaterial performance and the health of the host. Though this phenomenon has been studied for 50 years, our understanding is far from complete. The difficulty lies in the myriad of biomaterials designed for various applications along with the complexity of the adsorption event given the hundreds of proteins found in blood, their complex structure-function relationships and their potential interactions with one another. Protein-biomaterial interactions are too complex and the materials involved too varied to study on a case-by-case basis. Studies of protein adsorption to biomaterials with well characterized chemical modifications can be used to elucidate how molecular level modifications may impact protein adsorption, from which a generalized model of protein adsorption can be developed in addition to broad design strategies for future biomaterials. To this end, platelet poor human plasma was exposed to a variety of biomaterials, each with specific variations in their chemistry, and the composition of the adsorbed protein corona was evaluated using the highly sensitive and specific Western blotting method. Studies demonstrated that varying the level of carboxyl group substitution in a thermosensitive poly(ethylene glycol) and poly(ε-caprolactone) triblock copolymer hydrogel resulted in significantly different adsorbed proteomes. iii Increasing carboxyl substitution from 30% to 54% eliminated the presence of immunoglobulin G (IgG) and alpha1 antitrypsin while greatly increasing the presence of fibrinogen, prothrombin and antithrombin. In order to better understand the mechanism behind bioactive glass osseointegration and expand upon their successful clinical application, human plasma protein adsorption to bioactive glass 45S5 with varying crystallinities and/or manufacturing methods were evaluated. Increased crystallization did not affect the amount of protein adsorption, but did reduce the levels of deleterious proteins. Sol-gel manufacturing resulted in greater adsorption overall, particularly of fibrinogen and immunoglobulin G. An analysis of the proteomes adsorbed to poly(acrylic acid) nanoparticles containing various metal oxide cores indicated some significant variations in proteome composition including large differences in levels of complement factor 3 (C3), IgG, fibrinogen and fibronectin. The clotting response of human plasma in the presence of these particles tended to indicate varying degrees of inhibition, again depending upon the metal oxide core composition. A novel family of leucine-containing, short, marginally soluble elastin-like polypeptides (ELPs) were designed and produced in order to systematically examine the effects of chain length, guest amino acid chemistry and particle size on plasma protein adsorption as well as macrophage viability and phagocytosis. Given the unique combination of short lengths and hydrophobic guest amino acid, a novel purification method needed to be developed in order to successfully iv express and purify these ELPs which resulted in yields 10-20 times greater than reported yields for comparable constructs. A comprehensive, systematic examination of the leucine-containing ELP assembly and disassembly demonstrated previously unreported observations regarding ELP behaviour including how chain length, concentration and hydrophobicity may influence irreversible sub-micron particle formation as well as multi-micron, colloidally unstable aggregates. Hydrophobicity may have also influenced the zeta potential and packing density upon ELP assembly as well as particle stability upon dilution. Studies examining the adsorbed proteome as a function of ELP amino acid content, length and particle size found minor variations in plasma protein content and clotting response regardless of ELP composition or nanoparticle size. Moderate levels of adsorbed protein were found with the primary protein being albumin. These results suggest a generally favourable host response to these materials. ELP nanoparticles were found to have minimal impact on the viability of murine RAW 264.7 macrophage cells, regardless of the nanoparticle compositions or physical characteristics. Phagocytosis in this cell line was found to be mildly repressed in some instances as a function of nanoparticle diameter, amino acid hydrophobicity or ELP chain length. v The studies reported herein contribute to the development of a generalized model of biomaterial-protein adsorption and demonstrate the utility of short, marginally soluble ELPs as biocompatible nanoscale delivery vehicles.
This thesis is made available by the University of Alberta Libraries with permission of the copyright owner solely for the purpose of private, scholarly or scientific research. This thesis, or any portion thereof, may not otherwise be copied or reproduced without the written consent of the copyright owner, except to the extent permitted by Canadian copyright law.
Citation for previous publication
Yogasundaram H, Bahniuk MS, Singh HD, Aliabadi HM, Uludaǧ H, Unsworth LD. BSA nanoparticles for siRNA delivery: coating effects on nanoparticle properties, plasma protein adsorption, and in vitro siRNA delivery. International journal of biomaterials. 2012 Aug 7;2012.Bahniuk MS, Pirayesh H, Singh HD, Nychka JA, Unsworth LD. Bioactive glass 45S5 powders: effect of synthesis route and resultant surface chemistry and crystallinity on protein adsorption from human plasma. Biointerphases. 2012 Dec 1;7(1-4):1-5.Nikouei NS, Vakili MR, Bahniuk MS, Unsworth L, Akbari A, Wu J, Lavasanifar A. Thermoreversible hydrogels based on triblock copolymers of poly (ethylene glycol) and carboxyl functionalized poly (ε-caprolactone): the effect of carboxyl group substitution on the transition temperature and biocompatibility in plasma. Acta biomaterialia. 2015 Jan 15;12:81-92.Bahniuk MS, Alshememry AK, Unsworth LD. High-yield recombinant expression and purification of marginally soluble, short elastin-like polypeptides. BioTechniques. 2016 Dec;61(6):297-304.

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