Investigation of microbial cell deformability by filter cake compressibility using ultrafiltration membranes.

This study presents the investigation of deformability of various microbial cells in terms of filter cake compressibility during cake filtration using ultrafiltration membranes in dead-end mode. The examined microbial cells include mycoplasma, Gram-positive and Gram-negative bacteria, and Pseudomonas aeruginosa phage PP7. Polystyrene particles were used as an incompressible reference. The compressibility results were correlated to the deformability of a microbial cell, induced by its cell envelope. To determine the deformability of the different microbial cells under different process conditions, their cake resistance was measured under varying pressures from 10 to 250 kPa and temperatures from 2 to 35 °C. In addition, the influence of different culture media on the cell properties of Acholeplasma laidlawii and its behavior under different pressure and temperature was determined. The results of the pressure and temperature experiments revealed that Gram-positive S. epidermidis was found to be relatively stiff due to the thickness of the peptidoglycan layer, under different pressure and temperature conditions. No significant increase of the specific cake resistance of S. epidermidis could be determined. B. diminuta however showed a high deformation tendency when the pressure was increased indicating relatively soft cells. Mycoplasma A. laidlawii cells cultivated in three different media showed a different, but significant, effect of pressure and temperature.

Hybrid optimization of preparative chromatography for a ternary monoclonal antibody mixture.

In the purification of monoclonal antibodies, ion-exchange chromatography is typically used among the polishing steps to reduce the amount of product-related impurities such as aggregates and fragments, whilst simultaneously reducing HCP, residual Protein A and potential toxins and viruses. When the product-related impurities are difficult to separate from the products, the optimization of these chromatographic steps can be complex and laborious. In this paper, we optimize the polishing chromatography of a monoclonal antibody from a challenging ternary feed mixture by introducing a hybrid approach of the simplex method and a form of local optimization. To maximize the productivity of this preparative bind-and-elute cation-exchange chromatography, wide ranges of the three critical operational parameters-column loading, the initial salt concentration, and gradient slope-had to be considered. The hybrid optimization approach is shown to be extremely effective in dealing with this complex separation that was subject to multiple constraints based on yield, purity, and product breakthrough. Furthermore, it enabled the generation of a large knowledge space that was subsequently used to study the sensitivity of the objective function. Increased design space understanding was gained through the application of Monte Carlo simulations. Hence, this work proposes a powerful hybrid optimization method, applied to an industrially relevant process development challenge. The properties of this approach and the results and insights gained, make it perfectly suited for the rapid development of biotechnological unit operations during early-stage bioprocess development.