Improving glycosylation profiles and cell culture performance with a sensitive cell line in commercial manufacturing

A history of sporadically poor culture performance in the inoculum train and subsequent production cultures has been observed in a CHO cell line expressing a recombinant mAb X, resulting in insufficient titer for forward processing downstream. Through data analysis and experimentation in controlled bioreactors, this poor performance has been linked to seed train culture conditions- primarily seed train culture pH. Standard manufacturing practices for pH monitoring and control were modified to more precisely control culture pH to desired set point. Non-standard metrics (such as cell size and oxygen uptake rate) have been used to monitor seed train cultures. Evidence of the poor growth phenotype can also be found in elevated levels of binding immunoglobin protein (BiP) in the seed train culture cells. With seed train pH control improvements and enhanced monitoring in place, the poor culture performance has not been observed in recent commercial campaigns. In addition to reduced titer, product quality is also impacted when slower growth is observed in production culture, specifically out of specification high afucosylated species is observed. For one case of poor production performance that resulted in a high out of specification result for the afucosylated species, a review of equipment, cell age, raw material, adventitious agent testing, mechanical and process change records, and production media composition did not identify anything that could be linked to the slow growth observed. However, trend analysis revealed an abnormally high ammonium level in the early inoculum train stage. This case showed that minor deviations in the inoculum train cultures could result in poor performance in production cultures later, ultimately resulting in product quality failures. In order to reduce impact to product quality using this sensitive cell line, efforts to move the process into a more robust zone have been explored. Modified pH control strategies and impact to culture performance and product quality control will be discussed

Numerical modeling of unsaturated porous media as a two and three phase medium : a comparison.

A numerical simulation of the hydro-mechanical behaviour of an unsaturated porous medium is presented. The coupled hydro-mechanical model used is based on the continuum theory of mixtures and treats the unsaturated soil as a three phase porous medium (solid, liquid and gas). Principal variables are the solid deformation, the liquid pressure and the gas pressure. The two fluid phases are in motion and a non-linear pore pressure - saturation relation is used. The resulting system of equations is discretized in space using the finite element technique and in time by the - method. The comparison of numerical results and experimental test data shows that this hydro-mechanical model is capable to reproduce the principal phenomenon in the case of a hydric solicitation. The three phase formulation is compared to a simplified version considering only a two phase medium with static air phase. Merits and shortcomings of the two approaches are shown

Suction induced effects on the fabric of a structured soil

International audienceThis paper presents the mathematical modelling of the modification of the pore space geometry of a structured soil subjected to suction increase. Structured soil concepts are first introduced considering different fabric units, such as aggregates and fissures. The numerical modelling of the structural evolution is based on experimental test results in which the evolution of the structure of the samples subjected to different suctions is determined using the mercury intrusion porosimetry technique. From this information, the macro and micropore volume evolutions are determined. The results show that drying produces a reduction in the soil total porosity which mainly corresponds to a reduction of the macropore volume. Associated with this phenomenon, an increase in micropore volume is also observed. The proposed model divides pore size distribution into three pore classes (micropores, macropores and non-affected areas). Using the concept of a suction-influenced domain, the proposed model is able to reproduce the main observed fabric evolution between the saturated and dry states

Disturbed state concept for partially saturated soils.

A general elasto-plastic model for partially saturated soils is proposed, based on the disturbed state concept (DSC). This model is unified and simplified, includes both saturated and unsaturated states and is able to capture some typical features of partially saturated soils. The saturated state can be considered as a special case. Experimental results on a remolded sandy silt are used to calibrate the model

Comportement, calcul et dimensionnement des tranchées couvertes

La recherche effectuée sur les tranchées couvertes, ou tunnels exécutés à ciel ouvert, apporte une contribution théorique à la compréhension du comportement de ces ouvrages jusqu’à l’état limite ultime. Une méthode simplifiée inspirée de la méthode convergence-confinement est développée puis appliquée à plusieurs cas représentatifs. Cette méthode, basée sur le mécanisme de rupture de la structure, donne une solution approchée de l’état d’équilibre entre le sol et la structure. A ce titre, elle constitue un outil de conception car elle facilite la compréhension des mécanismes d’interaction sol-structure complexes caractéristiques de ces ouvrages. Le comportement du sol en interaction avec la structure est caractérisé typiquement par deux phases principales. La phase élastique est suivie par la plastification progressive de certaines zones du sol jusqu’à former un mécanisme plastique (phase plastique). Selon la géométrie et les matériaux, la structure profite alors plus ou moins favorablement de la contribution du sol à la reprise des charges. La conclusion principale de ce travail est l’existence de plusieurs régimes de comportement. Trois régimes principaux d’un intérêt pratique particulier, définis principalement par la phase de comportement du sol déterminante pour la structure, sont mis en évidence : sol élastique, sol élasto-plastique et sol complètement plastifié. L’identification du régime permet alors de concevoir efficacement la structure et renseigne sur les éléments importants à considérer lors du calcul et du dimensionnement. L’étude a également permis de montrer que la méthode dite aux modules de réaction n’est utilisable que dans certains cas bien définis. Dans les autres cas, la méthode aux éléments finis devrait être utilisée. Un nouveau format de sécurité compatible avec la nouvelle génération de normes SIA est proposé pour définir l’état limite ultime de la structure lors de l’utilisation de la méthode aux éléments finis. Ce format uniformise et clarifie la procédure de dimensionnement de ces structures. La recherche a aussi montré que la ductilité de la structure joue un rôle prépondérant pour garantir ou non l’exploitation des réserves de capacité offertes par le sol. Plusieurs limites de ductilité critiques pour les tranchées couvertes sont mises en évidence. Pour les tranchées couvertes de type cadre, la capacité de déformation de la dalle supérieure est très faible si aucun étrier n’est disposé dans les zones fortement sollicitées à l’effort tranchant. Pour les tranchées couvertes de type voûte, le phénomène d’éclatement du béton d’enrobage peut limiter la capacité de déformation de la structure. Des essais réalisés en laboratoire ont montré que la résistance à ce phénomène était influencée négativement par les déformations plastiques de l’armature et par la présence de joints de recouvrement. L’article 5.2.7.1 de la norme SIA 262 (2003) est jugée insuffisante. Des règles de vérification tirées des essais sont proposées. Le calcul de ces structures avec de grandes redistributions plastiques n’est alors possible que sous certaines conditions garantissant une ductilité suffisante

Water flow between soil aggregates

Aggregated soils are structured systems susceptible to non-uniform flow. The hydraulic properties depend on the aggregate fabric and the way the aggregates are assembled. We examined the hydraulic behavior of an aggregate packing. We focused on conditions when water mostly flows through the aggregates, leaving the inter-aggregate pore space air-filled. The aggregates were packed in 3mm thick slabs forming a quasi two-dimensional bedding. The larger aggregates were wetted with water and embedded in smaller aggregates equilibrated at a lower water content. The water exchange between wet and drier aggregates was monitored by neutron radiography. The three-dimensional arrangement of the aggregates was reconstructed by neutron tomography. The water flow turned out to be controlled by the contacts between aggregates, bottle-necks that slow down the flow. The bottle-neck effect is due to the narrow flow cross section of the contacts. The water exchange was simulated by considering the contact area between aggregates as the key parameter. In order to match the observed water flow, the contact area must be reduced by one to two orders of magnitude relative to that obtained from image analysis. The narrowness of the contacts is due to air-filled voids within the contact