bluuebunny/crossref_metadata_2025_split huggingface dataset:
High viable cell concentration fed‐batch cultures of hybridoma cells through on‐line nutrient feeding
Authors: Zhou Weichang , Rehm Jutta , Hu Wei‐Shou 
Published: December 2004

AbstractA hybridoma cell line was cultivated in fed‐batch cultures using a low‐protein, serum‐free medium. On‐line oxygen uptake rate (OUR) measurement was used to adjust the nutrient feeding rate based on glucose consumption, which was estimated on‐line using the stoichiometric relations between glucose and oxygen consumption. Through on‐line control of the nutrient feeding rate, not only sufficients were supplied for cell growth and antibody production, but also the concentrations of glucose and other important nutrients such as amino acids were maintained at low levels during the cell growth phase. During the cultivation, cell metabolism changed from high lactate production and low oxygen consumption to low lactate production and high oxygen consumption. As a result the accumulation of lactate was reduced and the growth phase was extended. In comparison with the batch cultures, in which cells reached a concentration of approximately 2 × 106 cells/mL, a very high concentration of 1.36 × 107 cells/mL with a high cell viability (>90%) was achieved in the fed‐batch culture. By considering the consumption of glucose and amino acids, as well as the production of cell mass, metabolites, and antibodies, a well‐closed material balance was established. Our results demonstrate the value of coupling on‐line OUR measurement and the stoichiometric realations for dynamic nutrient feeding in high cell concentration fed batch cultures. © 1995 John Wiley & Sons, Inc.
Preparation and in vitro analysis of microencapsulated genetically engineered E. coli DH5 cells for urea and ammonia removal
Authors: Prakash S. , Chang T. M. S. 
Published: December 2004

AbstractThis article describes a novel method of urea and ammonia removal using microencapsulated, genetically engineered Escherichia coli DH5 cells. Optimization of bacterial cell encapsulation was carried out. The optimal method consists of alginate 2.00% (w/v) at a flow rate of 0.0724 mL/min and a coaxial air flow rate of 2.00 L/min. This produces spherical, alginate‐poly‐L‐lysine‐alginate (APA) microcapsules of an average 500 ± 45 μm diameter. Increasing the concentration of alginate from 1.00% to 1.75% improves the quality of the microcapsules, while cell viability remains unaffected. The APA microcapsules are mechanically stable up to 210‐rpm agitation with no bacterial cell leakage. The in vitro performance of urea and ammonia removal by encapsulated bacteria is assessed. One hundred milligrams of bacterial cells in APA microcapsules, in their log phase state of growth, can lower 87.89 ± 2.25% of the plasma urea within 20 min and 99.99% in 30 min. The same amount of encapsulated bacteria can also lower ammonia from 975.14 ± 70.15 μM/L to 81.151 ± 7.37 μM/L in 30 min. There are no significant differences in depletion profiles by free and encapsulated bacteria for urea and ammonia removal. This novel approach using microencapsulated, genetically engineered E. coli cells is significantly more efficient than presently available methods of urea and ammonia removal. For instance, it is 30 times more efficient than the standard urease–ammonium adsorbent system. © 1995 John Wiley & Sons, Inc.
Growth kinetics of vitis vinifera cell suspension cultures: I. Shake flask cultures
Authors: Pépin Marie‐France , Archambault Jean , Chavarie Claude , Cormier François 
Published: December 2004

AbstractVitis vinifera cell suspension cultures carried out in shake flasks were closely examined for biomass growth and cell division in relation to carbohydrate, NH4, NO3PO4, and dissolved oxygen (DO)consumption. After inoculation, the oxygen uptake rate of the cultures measured on‐tine was observed to increase continuously to a maximum value of 3.8 mmol O2L−1h−1 at day 7 when cell division ceased and dissolved oxygen reached its lowest level of 17% air saturation. During this first phase of growth, the specific oxygen uptake rate remained constant at ∼0.6 mmol 02 O2 g−1 dw h−1or ∼2.2 μmol O2, (106 cells)−1 h−1 whereas dry biomass concentration increased exponentially from 1.5 to 6.0 g dw L−1. Thereafter, dry biomass concentration increased linearly to ∼14 g dw L−1 at day 14 following nitrate and carbohydrate uptake. During this second phase of growth, the biomass wet‐to‐dry weight ratio was found to increase in an inverse relationship with the estimated osmotic pressure of the culture medium. This corresponded to inflection points in the dry and wet biomass concentration and packed cell volume curves. Furthermore, growth and nutrient uptake results suggest that extracellular ammonium or phosphate ion availability may limit cell division. These findings indicate that cell division and biomass production of plant cell cultures may not always be completely associated, which suggests important new avenues to improve their productivity. © 1995 John Wiley & Sons, Inc.
Equilibria for the adsorption of antibiotics onto neutral polymeric sorbents: Experimental and modeling studies
Authors: Chaubal Mahesh V. , Payne Gregory F. , Reynolds Charles H. , Albright Robert L. 
Published: December 2004

AbstractThe objective of this work was to study the equilibria for adsorption of three antibiotics (penicillin V, tetracycline, and cephalosporin C) from water onto commercially available neutral polymeric sorbents. The pH was observed to be an important factor in adsorption as our results suggest that the neutral forms of penicillin V and cephalosporin C are preferentially adsorbed onto the neutral sorbents. Also, sorbent surface chemistry was observed to be important for adsorption, as the antibiotics adsorbed more favorably (both in terms of affinities and enthalpies) onto the aromatic sorbent as compared to the aliphatic ester sorbent. In addition to these thermodynamic measurements, molecular modeling studies and Monte Carlo simulations suggest that adsorption onto aromatic sorbents may involve specific interactions between the planar regions of the antibiotic molecules and the phenyl rings of the aromatic sorbent. The interaction energies predicted from Monte Carlo simulations were observed to provide qualitative agreement with experimentally determined adsorption affinities. © 1995 John Wiley & Sons, Inc.
Stability of transfectomas producing chimeric antibody against the pre‐S2 surface antigen of hepatitis B virus during a long‐term culture
Authors: Bae Sung Won , Hong Hyo Jeong , Lee Gyun Min 
Published: December 2004

AbstractTo design the scheme of large‐scale production of chimeric antibody for the postexposure prophylaxis of hepatitis B virus (HBV) infection, the stability of transfectomas (H69K‐1 and 6‐31) in regard to antibody production was examined during a long‐term, repeated fed‐batch culture without selection pressure using antibiotics. Although the H69K‐1 transfectoma was more stable than the 6‐31 transfectoma, both displayed gradual decreases in specific antibody productivity (qAb) for the first several weeks of cultivation. During this period, qAb was de‐creased by 40% to 50%. This loss of qAb was due mainly to the appearance of a nonproducing population (NP) of transfectoma, which was monitored throughout the culture by flow cytometry and the limiting dilution method. However, an NP did not overtake the culture and was balanced with a producing population (P) of transfectoma, resulting in stable antibody production. The subclones of NP obtained at the end of long‐term culture were further characterized by reverse transcription‐polymerase chain reaction assay of the heavy and light chain mRNA. All the subclones of NP derived from H69K‐1 transfectoma had only light chain mRNA. On the other hand, an NP in the 6‐31 transfectoma culture was heterogeneous. Some subclones of NP derived from 6‐31 transfectoma had only heavy chain mRNA and other subclones had only light chain mRNA. Taken together, the results obtained here suggest that selection pressure is necessary for a long‐term, continuous culture, because stable antibody production in a long‐term culture was achieved only after a significant loss of antibody productivity. Accordingly, a batch culture appears to be more appropriate for large‐scale chimeric antibody production without selection pressure. © 1995 John Wiley & Sons, Inc.
Interfacial properties of cell culture media with cell‐protecting additives
Authors: Michaels James D. , Nowak Jason E. , Mallik Ameet K. , Koczo Kalman , Wasan Darsh T. , Papoutsakis Eleftherios T. 
Published: December 2004

AbstractIn an effort to identify key rheological properties that contribute to cell protection against shear damage, we have measured surface shear and dilatationai viscosities, dynamic surface tension, foaminess, and foam stability for media containing cell‐protecting additives. In a companion article,18 we found that cell‐to‐bubble attachment was decreased in media containing Methocel, Pluronic F68, or polyvinyl alcohol (PVA). In medium containing polyethylene glycol (PEG) or potyvinyl‐pyrrolidone (PVP), attachment was increased. PEG, PVP, serum (FBS), and serum albumin (BSA) increased the surface viscosity of the air/medium surface (thus, producing a more rigid interface), whereas F68 and PVA lowered it greatly. Foaming experiments showed that Methocel, PEG, PVA, and F68 decreased the foam half‐life while FBS, BSA, and PVP were foam stabilizers. Interestingly, the foam stability of CHO cell suspensions decreased significantly for cell concentrations higher than ca. 2 × 106 cells/mL. Nonviable CHO cells reduced foam stability further. Dynamic surface tension values of the media tested were found significantly differentfrom their static surface tension values. The interfacial properties measured and the results presented in the companion study suggest that the additives that lower dynamic surface tension the most (Methocel, F68, and PVA) correlate well with reduced cell‐to‐bubble attachment, and thus, cell protection. Reduced dynamic surface tension with these additives implies faster surfactant adsorption, mobile interfaces, lower surface viscosity, and foam destabilization. Because PEG and PVP resulted in increased cell‐to‐bubble attachment and had different interfacial properties, a different mechanism (compared with Methocel, PVP, and F68) is apparently responsible for their protective effect. Finally, cell protection offered by FBS and BSA is attributed to the foam stabilization properties provided by these additives. © 1995 John Wiley & Sons Inc.
Towards the development of artificial endocrine tissues: 31P NMR spectroscopic studies of immunoisolated, insulin‐secreting AtT‐20 cells
Authors: Constantinidis I. , Sambanis A. 
Published: December 2004

AbstractTransformed, insulin‐secreting endocrine cells have been proposed as an alternative to islets for the development of a bioartificiat pancreas. With appropriate immunoprotection, such cells may be implanted without the need for patient immunosuppression. Use of continuous cell lines alleviates the cell availability limitation, but poses questions regarding thestability and biochemical and secretory function of the preparation, especially in the longterm. We have developed a bioreactor/perfusion system, compatible with a horizontal‐bore NMR instrument, that can maintain immunoprotected endocrine cells for prolonged periods of time. 31P NMR spectroscopy was used to study the bioenergetics of recombinant, insulin‐secreting mouse pituitary AtT‐20 cells entrapped as spheroids in calcium alginate/poly‐L‐lysine/alginate beads. NMR provided data verifying the macroscopic homogeneity within the bioreactor and allowing the evaluation of changes in cellular bioenergetics for a period of 70 days under different culture conditions. Levels of high‐energy phosphates changed slightly during the first 40 days of the experiment, then decreased considerably as cell death occurred. Rates of glucose consumption and insulin‐related peptide secretion also remained constant for 40 days and decreased rapidly thereafter. This study constitutes the beginning of an extensive quantitative analysis of the biochemistry of transformed endocrine cell lines in a sequestered, artificial tissue environment. © 1995 John Wiley & Sons Inc.
Cross‐flow ultrafiltration of proteins through asymmetric polysulfonic membranes: I. Retention curves and pore size distributions
Authors: Prádanos P. , Hernández A. 
Published: December 2004

AbstractFlux and retention of 0.1%w/w aqueous solutions of several proteins [lysozyme, pepsin, bovine serum albumin (BSA), lipase, and γ‐globulin] with molecular weights of 14.6, 36, 67, 801 and 150 kDa are studied when they are tangentially filtered, with transmembrane pressure differences until 1 MPa and circulation velocities in the re‐tentate loop from 0.04 to 1.98 m/s (laminar regime), through two asymmetric polysulfone commercial membranes (E‐100 with a nominal pore size of 0.01 μm and E‐500 with a nominal pore size of 0.04 μm). Results are analyzed with the film theory for the concentration‐polarization phenomenon, obtaining the mass transfer coefficient along with the apparent and true retention coefficients for the cell used, as a function of the feed circulation velocity and the molecular weight of the solute. The standard retention curves lead to pore size distributions differing from the nominal ones. These differences can be attributed to the modifications of the membranes when they are in operational conditions, probably due to protein adsorption. © 1995 John Wiley & Sons, Inc.
The effect of oscillating dissolved oxygen concentrations on the metabolism of a Spodoptera frugiperda IPLB‐Sf21‐AE clonal isolate
Authors: Rhiel Martin , Murhammer David W. 
Published: December 2004

AbstractThe effect of oscillating dissolved oxygen (DO) concentration on the metabolism of a clonal isolate of the Spodoptera frugiperda IPLB‐Sf21‐AE insect cell line was investigated. Specifically, the effect on cell growth, re‐ combinant protein synthesis, glucose and glutamine consumption, and lactate accumulation was determined. Prior to conducting the oscillating DO experiments, it was found that the DO concentration could be reduced to 15% air saturation without adversely affecting the growth rate. Under these conditions, glucose and glutamine became depleted as the maximum cell density was reached. The introduction of DO oscillations, that is, cycles consisting of 30 min at 15% DO followed by 30 min of anoxia, significantly altered cell metabolism, including inhibition of cell growth and recombinant protein synthesis. The effect of DO oscillations on glucose consumption was dependent on the experimental conditions. Glucose exhaustion occurred when the DO oscillations contained either an “apparent” anoxia period (nitrogen sparging discontinued upon reaching 0% DO) without pH control or a “true” anoxia period (nitrogen sparging continued throughout anoxia period) with pH control. Glucose consumption was significantly decreased, however, when the cells were exposed to a “true” anoxia period without pH control, that is, low pH inhibited glucose utilization. Glutamine uptake was not significantly affected by DO oscillations. Lactate only accumulated in the oscillating DO runs, a finding consistent with previous results demonstrating that significant lactate accumulation only occurs under DO‐limited conditions. © 1995 John Wiley & Sons, Inc.
A novel method to prepare multicellular spheroids from varied cell types
Authors: Yamazaki Manabu , Tsuchida Michiko , Kobayashi Ken‐yu , Takezawa Toshiaki , Mori Yuichi 
Published: December 2004

AbstractA simple method for preparing multicellular spheroids from varied cell types has been successfully developed by using a stepwise gradient surface in cell attachability or detachability. The surface was composed of poly‐N‐isopropylacrylamide (PNIPAAm), a temperature responsive polymer, as a cell detaching component, and collagen as a cell attaching component. The surface functions as a culture substratum at 37°C; then, when lowering the temperature of culture medium, the cells attached to it detach as a self‐supporting sheet. This is because PNIPAAm dissolves into the culture medium below the lower critical solution temperature (LCST; about 30°C), but it is insoluble above the LCST. The detached cell sheet forms a multicellular spheroid. The stepwise gradient surface which consisted of six different sectors was prepared by exposing a surface of the PNIPAAm‐collagen mixture to ultraviolet (UV) irradiation six times using a photomask, sliding the hole position in the photomask, and changing the energy of UV irradiation. This was because crosslinking of collagen depended on the energy of UV irradiation, then, cell attachability to and detachability from the surface were tightly controlled by changing the energy.The stepwise gradient surface allowed us to easily determine optimal surface conditions to obtain good cell attachment and detachment as a self‐supporting sheet from the surface to prepare multicellular spheroids. According to the evaluation of the attachability and detachability of 23 cell types, the optimal surface condition remarkably depended on each cell type. The detached cells under optimal surface conditions, including fibroblasts, osteoblastic cells, smooth muscle cells, and measangial cells, which were very difficult to form spherioids using conventional methods, were able to form multicellular spheroids. The results clearly demonstrate that the above‐described method for preparing multicellular spheroids can be applied to varied cell types. © 1995 John Wiley & Sons, Inc.
Comparison of hydrolysis and esterification behavior of Humicola lanuginosa and Rhizomucor miehei lipases in AOT‐stabilized water‐in‐oil microemulsions: I. Effect of pH and water content on reaction kinetics
Authors: Crooks Gavin E. , Rees Gareth D. , Robinson Brian H. , Svensson Martin , Stephenson G. Richard 
Published: December 2004

AbstractLipolase and Lipozyme are produced in large quantities (as a result of genetic engineering and overexpression) for the detergents market and provide a cheap source of highly active biocatalysts. Humicola lanuginosa lipase (HIL) and Rhizomucor miehei lipase (RmL) have been isolated in partially purified form from commercial preparations of Lipolase and Lipozyme, respectively. These lipases were solubilized in Aerosol‐OT (AOT)‐stabilized water‐in‐oil (w/o) microemulsions in n‐heptane. HIL and RmL activity in these microemulsions was assayed by spectrophotometric measurement of the initial rate of p‐nitophenyl butyrate hydrolysis, and by chromatographic determination of the initial rate of octyl decanoate synthesis from 1‐octanol and decanoic acid. The hydrolytic activity of HIL in microemulsions measured as a function of buffer pH prior to dispersal, followed a sigmoidal profile with the highest activities observed at alkaline pHs. This broadly matches the pH‐activity profile for tributyrin hydrolysis by Lipolase in an aqueous emulsion assay. The hydrolytic activity of RmL in the same microemulsions, measured as a function of pH, gave a bell‐shaped profile with a maximum activity at pH 7.5. Again, the observed pH‐activity profile was similar to that reported for a purified RmL in a tributyrin‐based aqueous emulsion assay. In contrast, the esterification activity exhibited by both HIL and RmL in AOT microemulsions over the available range pH 6.1 to 10.4, decreases as the pH increases, most likely reflecting the effect of substrate ionization. The dependence of the hydrolytic and condensation activity of HIL on R, the mole ratio of water to surfactant, were similar with both profiles exhibiting a maximum at R = 5. The hydrolytic and esterification activities of RmL followed similar R‐dependent profiles, but the profiles in this case exhibited a maximum at R = 10. The water activities at these R values were directly measured as 0.78 and 0.9, respectively. Measured water activities were unperturbed by the presence of lipase at the concentrations used in these studies. © 1995 John Wiley & Sons, Inc.
Mechanical disruption of Escherichia coli for plasmid recovery
Authors: Carlson Alfred , Signs Mark , Liermann Laura , Boor Robert , Jem K. Jim 
Published: December 2004

AbstractFive different mechanical cell disruption processes were evaluated as methods to extract plasmids from bacterial cells. The methods used were sonication, nebulization homogenization, microfluidization, and bead milling. The recovery yields of intact plasmids from the various methods were measured by quantitative gel electrophoresis. Bead milling and microfluidization were found to have the highest potential for large scale extraction with total intact recoveries of over 90% and around 50%, respectively. Other methods resulted in substantial plasmid degradation, with recoveries no greater than 20% of the total intact plasmid. © 1995 John Wiley & Sons, Inc.
Expanded bed affinity chromatography of dehydrogenases from bakers' yeast using dye–ligand perfluoropolymer supports
Authors: McCreath Graham E. , Chase Howard A. , Owen Ryan O. , Lowe Christopher R. 
Published: December 2004

AbstractMalate dehydrogenase (MDH) and glucose 6‐phosphate dehydrogenase (G6PDH) have been partially purified from preparations of homogenized yeast cells using Procion Yellow H‐E3G and Procion Red H‐E7B, respectively, immobilized on solid perfluoropolymer supports in an expanded bed. A series of pilot experiments were carried out in small packed beds using clarified homogenate to determine the optimal elution conditions for both MDH and G6PDH. Selective elution of MDH using NADH was effective but the yields obtained were dependent on the concentration of NADH used. Selective elution was found to be most effective when a low concentration of NaCl (0.1 M) was present. MDH could be recovered in 84% yield with a purification factor of 94 when this strategy was adopted. In the case of G6PDH, specific elution using NADP+ was successful in purifying G6PDH 178‐fold in 96% yield. The dynamic capacity of both affinity supports was estimated by frontal analysis, in an expanded bed with unclarified homogenate, and corresponded to 17 U MDH/mL of settled Procion Yellow H‐E3G perfluoropolymer support and 7.7 U H6PDH/mL of settled Procion Red H‐E7B perfluoropolymer support. Expanded bed affinity chromatography of MDH resulted in an eluted fraction containing 89% of the applied activity with a purification factor of 113. Expanded bed affinity chromatography of G6PDH resulted in an eluted fraction containing 84% of the applied activity with a purification factor of 172. With both enzymes, the overall recovery of enzyme activity was greater than 94%, showing that the expanded bed approach to purification was nondenaturing. © 1995 John Wiley & Sons, Inc.