L Day (l + day)

Distribution by Scientific Domains

Selected Abstracts

Cell Separator Operation within Temperature Ranges To Minimize Effects on Chinese Hamster Ovary Cell Perfusion Culture

Hans Drouin
A cell retention device that provides reliable high-separation efficiency with minimal negative effects on the cell culture is essential for robust perfusion culture processes. External separation devices generally expose cells to periodic variations in temperature, most commonly temperatures below 37 C, while the cells are outside the bioreactor. To examine this phenomenon, aliquots of ,5% of a CHO cell culture were exposed to 60 s cyclic variations of temperature simulating an acoustic separator environment. It was found that, for average exposure temperatures between 31.5 and 38.5 C, there were no significant impacts on the rates of growth, glucose consumption, or t-PA production, defining an acceptable range of operating temperatures. These results were subsequently confirmed in perfusion culture experiments for average exposure temperatures between 31.6 and 38.1 C. A 25,1 central composite factorial design experiment was then performed to systematically evaluate the effects of different operating variables on the inlet and outlet temperatures of a 10L acoustic separator. The power input, ambient temperature, as well as the perfusion and recycle flow rates significantly influenced the temperature, while the cell concentration did not. An empirical model was developed that predicted the temperature changes between the inlet and the outlet of the acoustic separator within 0.5 C. A series of perfusion experiments determined the ranges of the significant operational settings that maintained the acoustic separator inlet and outlet temperatures within the acceptable range. For example, these objectives were always met by using the manufacturer-recommended operational settings as long as the recirculation flow rate was maintained above 15 L day,1 and the ambient temperature was near 22 C. [source]

A Cyclical Semicontinuous Process for Production of Human ,1 -Antitrypsin Using Metabolically Induced Plant Cell Suspension Cultures

Melody M. Trexler
Transgenic rice suspension cultures were utilized to produce a human therapeutic protein, recombinant ,1 -antitrypsin (rAAT), in a cyclical, semicontinuous operation. Recombinant protein production was induced by removing the carbon source from the cell culture medium. The transgenic rice cells secreted the rAAT into the medium, and therefore medium exchanges could be performed for consecutive growth and protein expression phases. The process consisted of three cycles over a 25,28 day period, with growth phases lasting 4,6 days each and protein expression phases lasting 2.5,5 days each. Biomass and sugar concentrations, oxygen uptake rate, cell viability, culture pH, total extracellular protein, and active rAAT were measured throughout the cyclical process. The data profiles were reproducible between separate cyclical runs where, following each induction period, cell growth and viability could be reestablished once sucrose was added back to the culture. Volumetric productivities ranged from 3 to 12 mg active rAAT/(L day) for individual cycles with overall volumetric productivities of 4.5 and 7.7 mg active rAAT/(L day). [source]

Bioreactor Production of Human ,1 -Antitrypsin Using Metabolically Regulated Plant Cell Cultures

Melody M. Trexler
Transgenic rice cell cultures, capable of producing recombinant human ,1 -antitrypsin (rAAT), were scaled up from shake flasks to a 5-L bioreactor. The maximum specific growth rates (,max) observed from two bioreactor runs were 0.40 day,1 (doubling time of 1.7 days) and 0.47 day,1 (doubling time of 1.5 days), and the maximum specific oxygen uptake rates were 0.78 and 0.84 mmol O2/(g dw h). Using a metabolically regulated rice ,-amylase (RAmy3D) promoter, signal peptide, and terminator, sugar deprivation turned on rAAT expression, and rAAT was secreted into the culture medium. After 1 day of culture in sugar-free medium, there was still continued biomass growth, oxygen consumption, and viability. Extracellular concentrations of 51 and 40 mg active rAAT/L were reached 1.7 and 2.5 days, respectively, after induction in a sugar-free medium. Volumetric productivities for two batch cultures were 7.3 and 4.6 mg rAAT/(L day), and specific productivities were 3.2 and 1.6 mg rAAT/(g dw day). Several different molecular weight bands of immunoreactive rAAT were observed on immunoblots. [source]

Comparison of the microbial population dynamics and phylogenetic characterization of a CANOXIS reactor and a UASB reactor degrading trichloroethene

O. Tresse
Abstract Aims:, To understand the microbial ecology underlying trichloethene (TCE) degradation in a coupled anaerobic/aerobic single stage (CANOXIS) reactor oxygenated with hydrogen peroxide (H2O2) and in an upflow anaerobic sludge bed (UASB) reactor. Methods and Results:, The molecular study of the microbial population dynamics and a phylogenetic characterization were conducted using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). In both reactors, TCE had a toxic effect on two uncultured bacterial populations whereas oxygen favoured the growth of aerobic species belonging to Rhizobiaceae and Dechloromonas. No methanotrophic bacteria were detected when targeting 16S rRNA gene with universal primers. Alternatively, pmo gene encoding the particulate methane monooxygenase of Methylomonas sp. LW21 could be detected in the coupled reactor when H2O2 was supplied at 07 g O2 l day,1. Conclusions:,Methylomonas sp. LW21 that could be responsible for the aerobic degradation of the TCE by-products is not among the predominant bacterial populations in the coupled reactor. It seems to have been outcompeted by heterotrophic bacteria (Rhizobiaceae and Dechloromonas sp.) for oxygen. Significance and Impact of the Study:, The results obtained show the limitations of the coupled reactor examined in this study. Further investigations should focus on the operating conditions of this reactor in order to favour the growth of the methanotrophs. [source]

Dose selection and population pharmacokinetics of PEG-Intron in patients with chronic myelogenous leukaemia

Samir Gupta
Aims To assess the dose selection using population pharmacokinetics of Pegylated Intron-,2b (PEG-Intron) in patients with chronic myelogenous leukaemia (CML). Methods PEG-Intron 3,6 g kg,1 was administered subcutaneously once a week and blood samples were collected up to 48 weeks of treatment. A total of 624 samples collected from 137 patients were included in the analysis. Nonlinear mixed-effects modelling was used to analyse the sparsely sampled concentration data from a clinical efficacy trial. Covariates in the analysis included weight, sex, age, race, serum creatinine and estimated creatinine clearance (CLcr). Results The apparent clearance of PEG-Intron decreased after repeated dosing. The clearance at treatment week 4 was 42.3 l day,1 (patients with CLcr 120 ml min,1) with interpatient variability 30%. At treatment week 48, the clearance value was reduced to 69% of its week 4 value. CLcr, a composite variable calculated from body weight, sex, age and serum creatinine, had a small but statistically significant influence on the clearance of PEG-Intron. The clearance of PEG-Intron in patients with CML was 40% higher than that of hepatitis C virus-infected patients. Conclusion The dose of PEG-Intron 6.0 g kg,1 week,1 appeared appropriate in the treatment of patients with CML. [source]