Chemical and Biomolecular Engineering - Theses
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ItemProtein adsorption on particles: from particle engineering to understanding biomolecular corona formation( 2017)In biological fluids, biomolecules bind to particles, forming the biomolecular corona. This corona is generally described as a two-component system ‒ the “hard” and the “soft” corona. The adsorbed layer significantly influences the performance of the particles, both in vitro and in vivo. Thus, understanding and characterizing the biomolecular corona is essential. In this work, zwitterionic replica particles, consisting of poly(2-methacryloyloxy-ethyl phosphorylcholine) (PMPC), were synthesized by surface initiated-atom transfer radical polymerization. To understand factors that influence corona formation, the “hard” biomolecular corona formation on engineered particles in different biological milieu using microfluidic was examined. Key questions, such as the influence of flow, particle surface properties, incubation media, and incubation time were addressed. The data showed that dynamic incubation led to a more complex biomolecular corona, adsorption was suppressed on zwitterionic surfaces, and was enhanced after particle incubation in human blood, compared to human serum. An experimental approach that allowed for in situ monitoring of the “hard” and the “soft” protein corona was further introduced. The technique combined confocal laser scanning microscopy with microfluidics and allowed the study of the time-evolution of protein corona formation. The results showed that corona formation was kinetically divided into three different phases: Phase 1, proteins irreversibly and directly bound to the particle surface; Phase 2, irreversibly bound proteins interacting with pre-adsorbed proteins; Phase 3, reversibly bound “soft” corona proteins. To subsequently correlate the immune response to engineered particles and their “in situ” formed biomolecular corona, human whole blood assays were performed. Besides the usage of zwitterionic polymers, pre-adsorption of specific proteins to the particle’s surface is another commonly used technique to obtain low-fouling surfaces. To understand which parameters exactly, (I) the particle’s surface chemistry, (II) pre-adsorbed single proteins, or (III) the “in situ” formed biomolecular corona from human blood, predominantly trigger association with phagocytic cells, mass spectrometry analysis was combined with human whole blood assays. Mesoporous silica (MS) and zwitterionic PMPC particles were pre-coated with human serum albumin, immunoglobulin G, and complement protein C1q. Human whole blood assays revealed that cell–particle interactions were mainly dominated by the respective original surface chemistries of the particles, and less by the pre-adsorbed proteins. Furthermore, the compositions of the individual biomolecular coronas strongly differed between MS and PMPC particles, however remained similar for protein pre-coated systems. The presented work, allows understanding the consequences of particle design parameters and biological factors on (1) the corona composition, (2) its dynamic assembly and structural integrity, and (3) interactions with the human blood. The data might assist in the design of advanced drug delivery vehicles with improved circulation half-life due to suppressed protein adsorption.