Use of spectroscopic process analytical technology for rapid quality evaluation during preparation of CHO cell culture media.

Media preparation parameters contribute significantly to media quality, cell culture performance, productivity, and product quality. Establishing proper media preparation procedures is critical for ensuring a robust CHO cell culture process. Process analytical technology (PAT) enables unique ways to quantify assessments and improve media quality. Here, cell culture media were prepared under a wide range of temperatures (40-80°C) and pH (7.6-10.0). Media quality profiles were compared using three real-time PATs: Fourier-transform infrared (FTIR) spectroscopy, Raman spectroscopy, and excitation-emission matrix (EEM) spectroscopy. FTIR and Raman spectroscopies identified shifts in media quality under high preparation temperature (80°C) and at differing preparation pH which negatively impacted monoclonal antibody (mAb) production. In fed-batch processes for production of three different mAbs, viable cell density (VCD) and cell viability were mostly unaffected under all media preparation temperatures, while titer and cell specific productivity of mAb decreased when cultured in basal and feed media prepared at 80°C. High feed preparation pH alone was tolerated but cell growth and productivity profiles deviated from the control condition. Further, charge variants (main, acidic, basic species) and glycosylation (G0F, afucosylation, and high mannose) were examined. Statistically significant differences were observed for one or more of these quality attributes with any shifts in media preparation. In this study, we demonstrated strong associations between media preparation conditions and cell growth, productivity, and product quality. The rapid evaluation of media by PAT implementation enabled more comprehensive understanding of different parameters on media quality and consequential effects on CHO cell culture.

Optimization of adaptation parameters from adhesion cell culture in serum-containing media to suspension in chemically defined media by superlative box design.

A new design of experiments-superlative box design (SBD), was adopted to optimize the adaptation of Chinese hamster ovary cells from adhesion culture to serum-free suspension culture. It is a general trend to use a serum-free medium instead of a serum-containing medium. The advantage of serum-free medium (chemically defended) is that it does not contain unknown components and avoids safety issues. SBD requires fewer experiments while ensuring a sufficient number of experiments and uniformity in the distribution of experiments amongst all the factors. Six factors were considered in this experimental design with 43 runs plus three more repeating center runs. The cell line was adapted to serum-free media by gradually reducing serum, and from adherent to suspension by rotating at various speeds in a shake flask. Response surface methodology was applied to find the optimum condition. The optimized cell density reached 7.02 × 105 cells/mL, calculated by the quadratic model. Experiments validated the predicted cell adaptation with the maximum cell density. Three suspension runs were selected randomly to perform in the bioreactor to validate cell stability and production homogeneity. This study provides an efficient method to transfer adherent cells to suspension cells and is the first to successfully use SBD and establish a parameter quadratic optimization model.

Effects of different sodium salts and nitrogen sources on the production of 3-hydroxybutyrate and polyhydroxybutyrate by Burkholderia cepacia.

The effects of NaCl, Na2SO4, Na2HPO4, and Na3C6H5O7 on the production of 3-hydroxybutyrate, polyhydroxybutyrate, and by-products by Burkholderia cepacia. Proper addition of Na3C6H5O7 can significantly promote the production of 3-hydroxybutyric acid and polyhydroxybutyrate. The concentration, productivity, and yield of 3-hydroxybutyrate were increased by 48.2%, 55.6%, and 48.3% at 16 mM Na3C6H5O7. The increases of 80.1%, 47.1%, and 80.0% in the concentration, productivity, and yield of polyhydroxybutyrate were observed at 12 mM Na3C6H5O7. Na2SO4 and Na2HPO4 also have positive effects on the production capacity of 3-hydroxybutyrate and polyhydroxybutyrate within a certain range of concentration. NaCl is not conducive to the improvement of fermentation efficiency. Compared with a single nitrogen source, a mixed nitrogen source is more conducive to enhancing the production of 3-hydroxybutyrate and polyhydroxybutyrate.

Study on In-Situ Synthesis Process of Ti-Al Intermetallic Compound-Reinforced Al Matrix Composites.

In this study, ball-milled powder of Ti and Al was used to fabricate Ti-Al intermetallic compound-reinforced Al matrix composites by an in-situ reaction in cold-pressing sintering and hot-pressing sintering processes. The detailed microstructure of the Ti-Al intermetallic compound-reinforced Al composite was characterized by optical microscopy (OM), X-ray diffraction (XRD), energy dispersive spectrometry (EDS), and electron backscattered diffraction (EBSD). The results indicate that a typical core-shell-like structure forms in the reinforced particles. The shell is composed of a series of Ti-Al intermetallic compounds and has good bonding strength and compatibility with the Al matrix and Ti core. With cold-pressing sintering, the shell around the Ti core is closed, and the shell thickness increases as the milling time and holding time increase. With hot-pressing sintering, some radiating cracks emerge in the shell structure and provide paths for further diffusion of Ti and Al atoms. The Kirkendall effect, which is caused by the difference between the diffusion coefficients of Ti and Al, results in the formation of cavities and a reduction in density degree. When the quantity of the intermetallic compounds increases, the hardness of the composites increases and the plasticity decreases. Therefore, factors that affect the quantity of the reinforcements, such as the milling time and holding time, should be determined carefully to improve the comprehensive properties of the composites.