Determining Antibody Stability: Creation of Solid-Liquid Interfacial Effects within a High Shear Environment

Abstract

The purpose of this study was to assess the stability of protein formulations using a device designed to generate defined, quantifiable levels of shear in the presence of a solid-liquid interface. The device, based on a rotating disk, produced shear strain rates of up to 3.4 x 10(4) s(-1) (at 250 rps) and was designed to exclude air-liquid interfaces and enable temperature to be controlled. Computational fluid dynamics (CFD) was used to study the fluid flow patterns within the device and to determine the shear strain rate (s(-1)) at a range of disk speeds. The device was then used to study the effect on a monoclonal IgG4 of high levels of shear at the solid-liquid interface. Monomeric antibody concentration and aggregation of the protein in solution were monitored by gel permeation HPLC and turbidity at 350 nm. High shear strain rates were found to cause significant levels of protein aggregation and precipitation with reduction of protein monomer following first-order kinetics. Monomer reduction rate was determined for a range of disk speeds and found to have a nonlinear relationship with shear strain rate, indicating the importance of identifying and minimizing such environments during processing.

Study of robustness of filamentous bacteriophages for industrial applications

“…Such shear degradation profiles are consistent with those reported for other shear sensitive macromolecules (Biddlecombe et al, 2007;Zhang et al, 2007). Finally, the decay rate constants were found to increase in exponential proportion to the corresponding maximum energy dissipation rate, e (data not shown), according to the equation:…”

“…Shear-associated degradation by large-scale process equipment has been identified as a problem for several industrially relevant macromolecules, including plasmid DNA , whole antibodies (Biddlecombe et al, 2007) and protein precipitates (Boychyn et al, 2004). However, little equivalent information exists concerning the general response of filamentous bacteriophages to largescale processing.…”

“…In particular, a USD rotating disk shear device has been developed to predict the effects of turbulent fluid flow from large-scale pumps and continuous centrifuges (Boychyn et al, 2004;Hutchinson et al, 2006;Zhang et al, 2007). A solid base of small and large-scale verification experimentation exists, validating its use to determine patterns of shear-sensitive behavior for materials such as plasmid DNA, mammalian cell and antibody feeds (Biddlecombe et al, 2007;Hutchinson et al, 2006;Levy et al, 1999).…”

Aggregates in monoclonal antibody manufacturing processes

“…Stainless steel, a ubiquitous surface in bioprocessing, has been shown to cause aggregation of mAbs. Two different IgG4 mAbs were found to aggregate according to first-order kinetics when exposed to stainless steel under high shear conditions (Biddlecombe et al, 2007). In a different study, exposure of an IgG to stainless steel particulates caused the generation of much larger particles even when formulated with a non-ionic surfactant (Tyagi et al, 2009).…”

Micro biochemical engineering to accelerate the design of industrial‐scale downstream processes for biopharmaceutical proteins

“…An approach we have used and described as ''ultra scale-down'' is to seek out key effects which will dominate when scale changes are made in the processing of delicate biological materials. A central example of this kind of effect in bioprocessing is the sensitivity of biological cells, cell debris and protein precipitates to shear fields and of soluble proteins to shear in the presence of air-liquid interfaces or of solid-liquid interfaces (Biddlecombe et al, 2007). Where these effects apply strongly they determine performance and their evaluation at a very small scale can allow prediction of outcome at a large scale.…”