Thesis

Characterising immunoglobulin-polysorbate interactions

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Awarding institution
  • University of Strathclyde
Date of award
  • 2013
Thesis identifier
  • T13401
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Abstract
  • Therapeutic proteins, such as immunoglobulins, are typically formulated with polysorbates as stabilisers. However, the nature of the immunoglobulin-polysorbate interaction, particularly at the solid-liquid interface, is poorly characterised. This thesis presents an investigation of immunoglobulin (mAb-1)-surfactant interaction in bulk solution with particular focus on the interaction at the solid-liquid interface. It was first established using isothermal titration calorimetry that no specific binding interaction between mAb-1 and surfactant in solution takes place. Furthermore, circular dichroism and differential scanning calorimetry showed surfactant inclusion had no effect on mAb-1 native structure or thermal stability. The adsorption/desorption of mAb-1 and the effect of polysorbate was quantified in real-time by total internal reflection fluorescence. MAb-1 desorption was dependent on polysorbate concentration, fatty acid tail group and point of injection relative to mAb-1. MAb-1 adsorption to a hydrophobic surface was significantly less than to a hydrophilic surface. Concomitant conformational changes to mAb-1 were not apparent upon adsorption to a hydrophilic surface but a varying degree of β-sheet loss was observed upon adsorption to hydrophobic surfaces. This was corroborated by neutron reflectivity (NR) data which modelled a bilayer for mAb-1 adsorbed to a hydrophilic surface and a monolayer for mAb-1 adsorbed to a hydrophobic surface. These NR data suggested a range of mAb-1 orientations were adopted. This combination of orthogonal surface analytical techniques can build up a detailed molecular-level image of the adsorbed protein layer enabling rapid characterisation of protein surface adsorption which will improve bioprocess design and formulation.
Resource Type
DOI
Date Created
  • 2013
Former identifier
  • 989792

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