Glass microspargers as effective frit spargers in single use bioreactors.

For multiple-use bench scale and larger bioreactors, sintered stainless steel frit spargers are commonly used as microspargers. For bench-scale single-use bioreactors (SUBs), existing microspargers are sintered plastics, such as polyethylene. However, though plastics are readily sterilized by irradiation making them convenient for single use, these designs overlook surface energy properties of the materials of construction. For these sintered plastic spargers, forces at the water-air-surface interface cause bubble coalescence, leading to lower effective mass transfer, higher gas flow rates, and differing pCO2 profiles in cell culture. Alternative materials of construction were evaluated based on contact angle information and bubble formation observations. Sintered glass was chosen over thermoplastic polymers for higher surface wettability as described in the glass/water contact angle, its history as a commonly sintered material, and availability at costs suitable for single use applications. Glass sintered spargers and traditional stainless steel frit spargers were compared by porosity, bubble size, and kL a studies. Mass transfer (kL a) and cell culture performance equal or greater than a standard 20 µm stainless steel microsparger mass transfer efficiency was achieved by a glass frit sparger, of international porosity standard "P40" according to ISO 4793-80, which corresponds to a range of 16-40 µm.

Transient Electric Birefringence of Linear and Circular DNA: A Comparison of Kinetic Theory Predictions.

The use of monodisperse DNA and restriction enzyme modification allows the preparation of model samples of polymer rings and linear chains with a precision that is not possible by conventional synthetic routes. These samples allow direct comparisons of the predictions of polymer kinetic theories. Transient electric birefringence allows rapid imposition or forces (∼100 ns) and monitoring of conformational changes (∼0.7 µs). We determined the relaxation spectra of once-cut linear ΦX-174 (5386 bp), twice-cut linear ΦX-174 (2693 bp), and relaxed ring (5386 bp) ΦX-174 with transient electric birefringence. This allows comparison of the relaxation dynamics of a linear chain and a polymer loop with exactly the same contour length. The relaxed loop and the twice-cut DNA had the longest relaxation times of 1039 and 1076 µs, respectively. The loop was measured both in the native supercoiled state and in the relaxed state. We found the birefringence decayed in agreement with bead-spring (Rouse/Zimm) models for polymer dynamics determined by both a model-independent average relaxation time and deconvolution of the decay into a sum of exponentials. Deconvolution was performed by CONTIN and by the Padé-Laplace method. For both the relaxed ring and the linear fragments, we found the longer time constants τ1 and τ2 were discrete and had a spacing that was in agreement with the Rouse model without hydrodynamic interaction (τ1/τ2 = 0.25), which would be expected for a chain with 20 statistical segments. The excitation of higher relaxation modes could be observed as the electric field pulse length increased.

A comprehensive comparison of mixing, mass transfer, Chinese hamster ovary cell growth, and antibody production using Rushton turbine and marine impellers.

Large scale production of monoclonal antibodies has been accomplished using bioreactors with different length to diameter ratios, and diverse impeller and sparger designs. The differences in these physical attributes often result in dissimilar mass transfer, mechanical stresses due to turbulence and mixing inside the bioreactor that may lead to disparities in cell growth and antibody production. A rational analysis of impeller design parameters on cell growth, protein expression levels and subsequent antibody production is needed to understand such differences. The purpose of this study was to examine the impact of Rushton turbine and marine impeller designs on Chinese hamster ovary (CHO) cell growth and metabolism, and antibody production and quality. Experiments to evaluate mass transfer and mixing characteristics were conducted to determine if the nutrient requirements of the culture would be met. The analysis of mixing times indicated significant differences between marine and Rushton turbine impellers at the same power input per unit volume of liquid (P/V). However, no significant differences were observed between the two impellers at constant P/V with respect to oxygen and carbon dioxide mass transfer properties. Experiments were conducted with CHO cells to determine the impact of different flow patterns arising from the use of different impellers on cell growth, metabolism and antibody production. The analysis of cell culture data did not indicate any significant differences in any of the measured or calculated variables between marine and Rushton turbine impellers. More importantly, this study was able to demonstrate that the quality of the antibody was not altered with a change in the impeller geometry.