Rheology of clustering protein solutions

Abstract

High viscosity is a major challenge with protein therapeutics at extremely high concentrations. To overcome this obstacle, it is essential to understand the relationship between the concentration of a protein solution and its viscosity as a function of shear rate and temperature. Here, lysozyme is a model charged globular protein having both short-ranged attraction (SA) and long-ranged repulsion (LR) that promote the formation of dynamic clusters at high concentrations. We report viscosity measurements from a micro-capillary rheometer (using only several microliters of solution) over a wide range of lysozyme solution concentrations, shear rates, and temperatures. Solution structural relaxation dynamics are also probed by dynamic light scattering (DLS). As a result of lysozyme's SALR interactions, the viscosity increased dramatically across all shear rates with increasing concentration and decreasing temperature. While most of the solutions exhibited Newtonian behavior, shear thinning was exhibited at the highest concentration (480 g/l) and lowest temperatures at shear rates above approximately 10(4 )s(-1). The onset shear rate for thinning and a structural relaxation rate estimated from a slow-mode measured by DLS are compared. These measurements provide insights into the properties of protein solutions and their microscopic structural origins.

Dynamics of proteins in solution

“…Other non-neutron based techniques such as DLS (Bolañnos-García et al, 1998;Li et al, 2011;Arzenšek et al, 2012;Soraruf et al, 2014;Maes et al, 2015;Bauer et al, 2016), DLS combined with Raman spectroscopy (Lewis et al, 2014) or with NMR (Poznański et al, 2005), fluorescence spectroscopy (Nath et al, 2010;Roberti et al, 2011;Nath and Rhoades, 2013) and rheology (Dharmaraj et al, 2016), as well as simulations (Bratko et al, 2007) can be used to study the dynamics of systems with aggregating proteins. One of the strengths of the combination of such techniques with neutron scattering related to the different accessible times, is the potential of inferring information on the type and lifetime of clusters (as in Liu et al, 2010) and aggregates on wide concentration ranges, as well as on the kinetics of the process.…”

Pressure drop in liquid chromatography

“…However, this is not necessarily the case for solutions of macromolecules. While most of the protein solutions behaves like Newtonian fluids in a concentration range encountered in chromatography , molecules like DNA express shear tinning already in concentration of several mg/ml . As a consequence at sufficiently high linear velocity viscosity decreases due to orientation of DNA molecules in flow direction what might cause that pressure drop increases slower than the flow‐rate increase – a negative deviation from linearity.…”

The NISTmAb Reference Material 8671 lifecycle management and quality plan

“…The current catalog of NIST materials support a broad range of disciplines and measurements including amount of substance, dimensional metrology, electricity and magnetism, mass and related quantities, time and frequency, among others [ 23 ]. Biomanufacturing is an expanding priority area for NIST as demonstrated by numerous recent publications on emerging scientific technologies including high resolution protein structure measurements [ 4 – 7 ], mass spectral libraries [ 24 ], rheology and behavior of high concentration protein solutions [ 25 ], protein particulates [ 26 , 27 ], and cell therapies [ 28 ]. A key component of NIST’s Biomanufacturing Initiative is the development of industry-focused RMs designed to enable more accurate and confident characterization of key attributes linked to product safety and efficacy.…”