Improvement of the ITC binding analysis methodology for the unspecific interaction between serum proteins and polymer brushes grafted to nanoparticles

• Main Author: Leitner, Nikolaus Simon
• Other Authors: -
• Format: Thesis-Doctor of Philosophy
• Language: English
• Published: Nanyang Technological University 2023
• Subjects: Engineering::Materials
• Online Access: https://hdl.handle.net/10356/165155
• Institution: Nanyang Technological University

The use of nanoparticles in consumer and industrial applications is widespread and proliferates annually. Their tunable physicochemical properties improved many products and processes. The utilization of nanoparticles is mostly unregulated or underregulated, which arouses urgent questions about the direct and long-term effects on humans and the environment. Such questions are insufficiently answered because analysis methods capable of probing the weak interactions between proteins and particles have their own limitations. These analytical obstacles can also curb the development of new nanoparticle formulations. Other reasons for these uncertainties regarding the biosafety of nanoparticles lie in the complex interaction between nanoparticles with biomolecules, cells, and organs. Biomedical nanoparticles are usually injected into the bloodstream or directly into the target tissue. Due to the particle's attractive force, biomolecules form a strongly bound “hard corona” and a highly dynamic “soft corona“. Softand hard corona are forming the “protein corona” , which composition is heavily influenced by the physicochemical properties of the nanoparticle. The protein corona is responsible for the cell-nanoparticle interaction and, therefore, also for nanomaterials' ultimate biological fate. Unfortunately, current available analytical methods either probe the protein corona with significant limitations only. This thesis addresses the complex nature of the protein corona and the limitation and strengths of commonly used analytical methods. This work focuses on measuring affinity, stoichiometry, and thermodynamic parameters of the weak interaction between thermoresponsive polymer brushes densely grafted on small superparamagnetic iron oxide nanoparticles (SPIONs) and blood proteins with the rarely used isothermal titration calorimetry (ITC) method. This approach showed that lysozyme and transferrin are not interacting with certainty with the used SPIONs. On the other hand, the binding- and thermodynamic parameters of the interaction of albumin and immunoglobulin G with SPIONs could be determined at physiological protein concentrations. Additionally, ITC was used to compare the interaction of albumin with SPIONs which have a shell consisting of linear or cyclic polymers. In contrast to linear polymer shells, particles with cyclic brushes did not show interaction with albumin. Furthermore, the sequestering of toxic heavy metal ions by a novel polymer was quantified with ITC. The following work aims to improve the methodology of ITC for the analysis of the interaction between core-shell nanoparticles with biomolecules and ions. It compares ITC to other analytical methods generally accepted for binding studies and could be used as a starting point for new studies on the protein corona.