Characterization of a single-use stirred-tank bioreactor vessel for microcarrier-based adherent cell culture processes using experimental and computational fluid dynamics studies

Renewed interest in microcarrier-based processes for the large-scale culture of adherent cells for vaccine and cell therapy applications drives the need for effective, high-throughput, single-use, process development tools that can be translated successfully into industrial-scale systems. The automated ambr250® platform is one such technology, operating at a volume between 100 – 250mL and which is both high-throughput and single-use. The ambr250 has demonstrated significant success for suspension-based mammalian cell culture applications. However, no studies have been reported investigating microcarrier-based processes for the culture of adherent cells. With any cell culture process, the fluid dynamics characteristics of the bioreactor must be sufficiently well understood to enable successful scale-up to larger scale bioreactors. With microcarriers, there is an additional challenge as the fluid dynamics must take into account the presence of the particulate solid phase. A critical aspect for cell cultivation on microcarriers is the minimum agitator speed required to achieve complete microcarrier suspension, NJS. Under these conditions, the surface area of the attached cells is available for transfer of nutrients (including oxygen) to the cells and metabolites from them, whilst higher speeds hardly increase these transport processes and may lead to damaging fluid dynamic stresses being generated1. This suspension condition can be studied experimentally if equipment is specially modified to make easy visual observation of the two-phase flow in the bioreactor which during actual culture is very difficult. Therefore, it is extremely beneficial to both measure NJS and then to compare the measured values with predictions based on computational fluid dynamics (CFD) in order to validate the latter. Once validated, CFD modelling is a very useful tool for analysing flow patterns, mixing time, mean and local specific energy dissipation rates and other parameters important for scale up in order to optimise the overall bioreactor geometry. In addition to the above fluid dynamic aspects, cell culture studies was also performed in parallel to analyse the cell growth at and around the minimum speed for microcarrrier suspension, NJS and the results were compared to the culture performance in well-characterised traditional spinner flask bioreactors2.The CFD and experimental results with the single-use ambr250 bioreactor will be discussed in detail along with their scale-up implications. References 1. Nienow, A. W., Coopman, K., Heathman, T. R. J., Rafiq, Q. A. and C. J. Hewitt (2016). “Bioreactor Engineering Fundamentals for Stem Cell Manufacturing”. In: “Stem Cell Manufacturing”, (Eds. J.M.S. Cabral, C.L. de Silva, L. G. Chase and M. M. Diogo), Elsevier Science, Cambridge, USA; Chapter 3, pp 43 – 76. 2. Rafiq, Q. A., Brosnan, K. M., Coopman, K., Nienow, A. W. and Hewitt, C.J. (2013) Culture of Human Mesenchymal Stem Cells on Microcarriers in a 5 L Stirred-Tank Bioreactor. (with Q. A. Rafiq, K. M. Brosnan, K

Characterization of a single-use stirred-tank bioreactor vessel for microcarrier-based adherent cell culture processes using experimental and computational fluid dynamics studies

Renewed interest in microcarrier-based processes for the large-scale culture of adherent cells for vaccine and cell therapy applications drives the need for effective, high-throughput, single-use, process development tools that can be translated successfully into industrial-scale systems. The automated ambr250® platform is one such technology, operating at a volume between 100 – 250mL and which is both high-throughput and single-use. The ambr250 has demonstrated significant success for suspension-based mammalian cell culture applications. However, no studies have been reported investigating microcarrier-based processes for the culture of adherent cells. With any cell culture process, the fluid dynamics characteristics of the bioreactor must be sufficiently well understood to enable successful scale-up to larger scale bioreactors. With microcarriers, there is an additional challenge as the fluid dynamics must take into account the presence of the particulate solid phase. A critical aspect for cell cultivation on microcarriers is the minimum agitator speed required to achieve complete microcarrier suspension, NJS. Under these conditions, the surface area of the attached cells is available for transfer of nutrients (including oxygen) to the cells and metabolites from them, whilst higher speeds hardly increase these transport processes and may lead to damaging fluid dynamic stresses being generated1. This suspension condition can be studied experimentally if equipment is specially modified to make easy visual observation of the two-phase flow in the bioreactor which during actual culture is very difficult. Therefore, it is extremely beneficial to both measure NJS and then to compare the measured values with predictions based on computational fluid dynamics (CFD) in order to validate the latter. Once validated, CFD modelling is a very useful tool for analysing flow patterns, mixing time, mean and local specific energy dissipation rates and other parameters important for scale up in order to optimise the overall bioreactor geometry. In addition to the above fluid dynamic aspects, cell culture studies was also performed in parallel to analyse the cell growth at and around the minimum speed for microcarrrier suspension, NJS and the results were compared to the culture performance in well-characterised traditional spinner flask bioreactors2.The CFD and experimental results with the single-use ambr250 bioreactor will be discussed in detail along with their scale-up implications. References 1. Nienow, A. W., Coopman, K., Heathman, T. R. J., Rafiq, Q. A. and C. J. Hewitt (2016). “Bioreactor Engineering Fundamentals for Stem Cell Manufacturing”. In: “Stem Cell Manufacturing”, (Eds. J.M.S. Cabral, C.L. de Silva, L. G. Chase and M. M. Diogo), Elsevier Science, Cambridge, USA; Chapter 3, pp 43 – 76. 2. Rafiq, Q. A., Brosnan, K. M., Coopman, K., Nienow, A. W. and Hewitt, C.J. (2013) Culture of Human Mesenchymal Stem Cells on Microcarriers in a 5 L Stirred-Tank Bioreactor. (with Q. A. Rafiq, K. M. Brosnan, K

Experimental and Computational Fluid Dynamics study of microcarrier suspension during the cultivation of Mesenchymal Stem Cells in an ambr250 bioreactor

The ambr250 unit is a fully automated disposable 100-250 ml bioreactor for R&D that has been developed by TAP Biosystems, now part of Sartorius-Stedim, widely used for scale down and scale up modelling studies. Recently, mesenchymal stem cells (MSCs) have become strong candidates for cell-based therapies based on in vitro growth on microcarriers in stirred bioreactors. However, to fully realize the MSCs potential, a number of key processing issues need to be addressed because of the huge number of cells that are required. Thus, the fluid dynamics characteristics of the stirred ambr250 bioreactor must be sufficiently well understood to enable scale-up to larger bioreactors to be efficiently accomplished particularly because of the special issues arising from the presence of the particulate solid phase. One of the most critical aspects for MSC cultivation on microcarriers is the minimum agitator speed required to achieve complete microcarriers suspension, NJS. Under these conditions, the surface area of all the attached cells is available for transfer of nutrients (including oxygen) to the cells and metabolites from them, whilst higher speeds hardly increase these transport processes and may lead to damaging fluid dynamic stresses being generated1. This suspension condition can be studied experimentally if equipment is specially modified to make easy visual observation of the two-phase flow in the bioreactor but during actual growth that is very difficult. Therefore, it is extremely beneficial to both measure NJS and compare the measured values with predictions based on computational fluid dynamics (CFD) for validation. Once validated, then CFD is a very useful tool for analyzing flow patterns, mixing time, mean and local specific energy dissipation rates and other parameters important for scale up. In this work we examined the fluid dynamics of the two-phase particle-liquid system with an experimental analysis and a CFD simulation using a lattice-Boltzmann base software and particle tracking of an ambr250 vessel at different stirring conditions. Cell culture was also performed in parallel to analyse the cell growth at and around NJS and the results were compared to the performance in a spinner flask bioreactor. The CFD and experimental results will be discussed in detail along with their scale-up implications. References 1) Nienow, A. W., Coopman, K., Heathman, T. R. J., Rafiq, Q. A. and C. J. Hewitt (2016). “Bioreactor Engineering Fundamentals for Stem Cell Manufacturing”. In: “Stem Cell Manufacturing”, (Eds. J.M.S. Cabral, C.L. de Silva, L. G. Chase and M. M. Diogo), Elsevier Science, Cambridge, USA; Chapter 3, pp 43 – 76

A Test of Multisession Automatic Action Tendency Retraining to Reduce Alcohol Consumption Among Young Adults in the Context of a Human Laboratory Paradigm.

BACKGROUND: Young adult heavy drinking is an important public health concern. Current interventions have efficacy but with only modest effects, and thus, novel interventions are needed. In prior studies, heavy drinkers, including young adults, have demonstrated stronger automatically triggered approach tendencies to alcohol-related stimuli than lighter drinkers. Automatic action tendency retraining has been developed to correct this tendency and consequently reduce alcohol consumption. This study is the first to test multiple iterations of automatic action tendency retraining, followed by laboratory alcohol self-administration. METHODS: A total of 72 nontreatment-seeking, heavy drinking young adults ages 21 to 25 were randomized to automatic action tendency retraining or a control condition (i.e., sham training ). Of these, 69 (54% male) completed 4 iterations of retraining or the control condition over 5 days with an alcohol drinking session on Day 5. Self-administration was conducted according to a human laboratory paradigm designed to model individual differences in impaired control (i.e., difficulty adhering to limits on alcohol consumption). RESULTS: Automatic action tendency retraining was not associated with greater reduction in alcohol approach tendency or less alcohol self-administration than the control condition. The laboratory paradigm was probably sufficiently sensitive to detect an effect of an experimental manipulation given the range of self-administration behavior observed, both in terms of number of alcoholic and nonalcoholic drinks and measures of drinking topography. CONCLUSIONS: Automatic action tendency retraining was ineffective among heavy drinking young adults without motivation to change their drinking. Details of the retraining procedure may have contributed to the lack of a significant effect. Despite null primary findings, the impaired control laboratory paradigm is a valid laboratory-based measure of young adult alcohol consumption that provides the opportunity to observe drinking topography and self-administration of nonalcoholic beverages (i.e., protective behavioral strategies directly related to alcohol use)

Experimental and computational fluid dynamics studies of adherent cells on microcarriers in an ambr® 250 bioreactor

Interest for microcarrier-based processes for the large-scale culture of adherent cells has recently grow, due to possible application in vaccine and cell therapy. This opportunity drives the need for effective, high-throughput, single-use, process development tools that can be translated successfully into industrial-scale systems. The automated ambr® 250 platform is one such technology, operating at a volume between 100 – 250mL, both high-throughput and single-use. The ambr250 has demonstrated significant success for suspension-based mammalian cell culture applications. However, additional investigations need to be performed on microcarrier-based processes for the culture of adherent cells. The fluid dynamics characteristics of the bioreactor must be sufficiently well understood to enable successful scale-up to larger scale bioreactors. Physical parameters such as fluid velocity, power number and shear stress are important for any cell culture. With microcarriers, there is an additional challenge as the fluid dynamics must take into account the presence of the particulate solid phase. A critical aspect for cell cultivation on microcarriers is the minimum agitator speed required to achieve complete microcarrier suspension, NJS. Under these conditions, the surface area of the attached cells is available for transfer of nutrients (including oxygen) to the cells and metabolites from them, whilst higher speeds hardly increase these transport processes and may lead to damaging fluid dynamic stresses being generated. It is also extremely beneficial to predict the flow dynamics of the stirred tank based on computational fluid dynamics (CFD). Once validated, CFD modelling is a very useful tool for analysing flow patterns, mixing time, mean and local specific energy dissipation rates, shear stress, and other parameters important for scale up in order to optimise the overall bioreactor geometry. In addition to the above fluid dynamic aspects, cell culture studies was also performed in parallel to analyse the cell growth at and around the minimum speed for microcarrier suspension, NJS. The CFD and experimental results with the single-use ambr250 bioreactor will be discussed in detail during the final presentation

Internet self-efficacy does not predict student use of Internet-mediated educational technology

Two studies tested the hypothesis that use of learning technologies among undergraduate psychology students was associated with higher Internet self-efficacy (ISE). In Study 1, the ISE scores of 86 students were found not to be associated with either attitudes towards, or measured use of, blogs and wikis as part of an IT skills course. ISE was associated with time spent online, and positive attitudes to wikis were associated with higher use. Study 2 measured 163 students’ ISE scores at the beginning and end of the same course. ISE was again not correlated with attitudes towards, or actual measured use of, learning technologies used in the course. However, ISE was shown to increase during the course. Positive attitudes towards wikis and discussion boards were associated with higher use of each. Overall, ISE scores did not influence measured use of a Virtual Learning Environment (VLE, including blogs, wikis and a discussion board), or attitudes towards those technologies. This implies that while ISE is linked to aspects of online behaviour (time spent online) and can be modified by online activity or training, it does not predict student use of educational Internet technologies

The confluence model: Parameter estimation for six divergent data sets on family factors and intelligence

Theoretical predictions, based on the confluence model, were made for data from six national surveys of intellectual performance. All six surveys relate intellectual performance scores to family configuration variables. Widely divergent patterns of relationships between the major family factors, such as birth order and family size, and intellectual performance scores characterize the six data sets. Nevertheless, the confluence model was capable of accurate prediction in all cases when all three parameters were estimated. Little accuracy was lost using only two parameters. Variations in the estimated parameter values that reflected the variations in patterns of effects could be meaningfully interpreted for their psychological significance.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/23137/1/0000061.pd

Revisiting Consumer Ethnocentrism: Review, Reconceptualization, and Empirical Testing

Prior research has suggested that many consumers prefer domestic to foreign products, even when the quality is lower and the price is higher. Such bias is attributed to consumer ethnocentrism. This study critically examines the current conceptualizations of consumer ethnocentrism and proposes an extension of its conceptual boundaries and measurement. It determines that consumer ethnocentrism is a multidimensional construct that encompasses five dimensions: prosociality, cognition, insecurity, reflexiveness, and habituation. Empirical evidence from the United Kingdom and the United States demonstrates that the extended measurement instrument better predicts consumers' preferences for local brands at the expense of foreign brands

Mouth Rinsing With Carbohydrate Solutions at the Postprandial State Fail to Improve Performance During Simulated Cycling Time Trials.

Mouth-rinsing with carbohydrate solutions during cycling time trials results in performance enhancements, however the majority of studies have utilised ∼6% carbohydrate solutions. Therefore, the purpose of this study was to compare the effectiveness of mouth-rinsing with 4%, 6%, and 8% carbohydrate (CHO) solutions on 1-h simulated cycling time trial performance. On four occasions, seven trained male cyclists completed at the postprandial period, a set amount of work as fast as possible in a randomised, counterbalanced order. The subjects mouth rinsed for 5-s, upon completion of each 12.5% of the trial, with 25 mL of a non-CHO placebo, 4%, 6%, and 8% CHO solutions. No additional fluids were consumed during the time trial. Heart rate (HR), ratings of perceived exertion (RPE), thirst (TH) and subjective feelings (SF) were recorded after each rinse. Further, blood samples were drawn every 25% of the trial to measure blood glucose (BG) and blood lactate (BG) concentrations, whilst whole body carbohydrate oxidation was monitored continuously. Time to completion was not significant between conditions with the placebo, 4%, 6%, and 8% conditions completing the trials in 62.0 ± 3.0, 62.8 ± 4.0, 63.4 ± 3.4, and 63 ±4.0 minutes respectively. There were no significant differences between conditions in any of the variables mentioned above however significant time effects were observed for HR, RPE, TH, and SF. Post-hoc analysis showed that TH and SF of subjects in the CHO conditions but not in the placebo were significantly increased by completion of the time trial. In conclusion, mouth-rinsing with CHO solutions did not impact 1-h cycling performance in the postprandial period and in the absence of fluid intake. Our findings suggest that there is scope for further research to explore the activation regions of the brain and whether they are receptive to CHO dose, before specific recommendations for athletic populations are established. Consequently mouth-rinsing as a practical strategy for coaches and athletes is questionable under specific conditions and should be carefully considered before its inclusion. The emphasis should be focused on appropriate dietary and fluid strategies during training and competition