摘要:

AbstractTo investigate the effects of factors secreted by different cell lines on human monoclonal antibody (MAb) integrity, 600 mg of a human MAb, which specifically binds to human erythrocytes, were produced in a perfusion process. After purification by protein A affinity chromatography, the MAb was used for integrity testing in supernatants of several cell lines to investigate their potential to degrade the antibody in the extracellular environment. One insect cell line (IPLB‐SF‐21 AE) and four mammalian cell lines [CHO K1, BHK‐21 (C13), C1271, P3‐X63‐Ag8.653], all of them commonly used for the production of recombinant proteins, and the human–human–mouse heterohybridoma cell line itself (H‐CB‐hahE), were adapted to serum‐free culture media. For integrity testing all cell lines were cultivated in spinner flasks using serum‐free media supplemented with 30 μg mL−1of purified MAb. MAb integrity was assayed by SDS polyacrylamide gel electrophoresis (SDS‐PAGE), isoelectric focusing, both followed by Western blotting, and an antigen binding assay. None of the mammalian cells showed any detectable effects on antibody stability and integrity during exponential growth, whereas isoelectric focusing of monoclonal antibody taken from IPLB‐SF‐21 AE culture supernatants revealed a new band indicating a partial modification of the MAb by secreted factors of these cells. This observation did not correlate with the total proteolytic activity, which was measured in all supernatants and found to be lowest in the insest cell cultures. For mammalian cell cultures, it could be concluded from these findings that shifts of the antibody microheterogeneity pattern, which can be found normally as a result of variations in different production parameters, are not caused by extracellular factors once the product has been secreted into the supernatant. In addition to their well‐known advantages in posttranslational modifications (e.g., formation of complex typeN‐glycans), mammalian cells appear to be more suitable as expression systems for human monoclonal antibodies to be used in vivo when compared with baculovirus‐infected insec

ISSN:0006-3592    

DOI:10.1002/bit.260450202

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractA novel type of bioreactor for waste gas treatment has been designed. The reactor contains a microporous hydrophobic membrane to create a large interface between the waste gas and the aqueous phase. To test the new reactor, propene was chosen because of its high air/water partition coefficient, which causes a low water concentration and hampers its removal from air. Propene transfer from air to a suspension of propene‐utilizingXanthobacterPy2 cells in the membrane bioreactor proved to be controlled by mass transfer in the liquid phase. The resistance of the membrane was negligible. Simulated propene transfer rates agreed well with the experimental data. A stable biofilm ofXanthobacterPy2 developed on the membrane during prolonged operation. The propene flux into the biofilm was 1 × 10−6mol m−2s−1at a propene concentration of 9.3 × 10−2mol m−3in the gas phase. © 1995 John W

ISSN:0006-3592    

DOI:10.1002/bit.260450203

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractA molecular assembly in which a conjugated polymer is interfaced with a photodynamic protein is described. The conjugated polymer, functionalized with biotion, is designed such that it can be physisorbed on or chemically grown off a glass surface. The streptavidin‐derivatized protein is immobilized on the biotinylated polymer matrix through the strong biotin–streptavidin interactions. The assembly, built on the surface of an optical fiber or on the inside walls of a glass capillary, forms an integral part of a biosensor for the detection of environmental pollutants such as organophosphorus‐based insecticides. The Protein in the system can be replaced by any biological macromolecule of interest. We study one specific case, the enzyme alkaline phosphatase. The enzyme catalyzes a reaction producing an intermediate compound that chemiluminesces, and the chemiluminescence singnal is monitored to detect and quantify insecticides such as paraoxon and methyl parathion. Preliminary results indicate ppb level detection with response time less than 1 minute. © 1995 John Wiley&Son

ISSN:0006-3592    

DOI:10.1002/bit.260450204

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractAn on‐line calibration procedure for application in continuous monitoring systems has been developed. Control of the calibration value and recalibration on‐line during monitoring is possible without having to disrupt the sample withdrawal. The calibration procedure has been applied and evaluated in a continuous biosensor system based on the detection of oxygen depletion during enzymatic substrate conversion by immobilized oxidases. Evaluation included on‐line calibration during continuous measurements of glucose and lactate in bovine blood samples. Calibration of the complete system consisting of a sampling device, a sample handling step, a biocatalytic step, a detection step, and a data processing unit is performed by the on‐line addition of a calibration solution to a blank sample which is fed through the system. The calibration cycle is completed within 5.5 min. When recalibration is carried out during monitoring, the calibration solution is added to the sample, instead of to a blank sample, and the increase in outlet singl is registered. The major advantage of this internal standard principle is that the calibration solution is fed through the whole system according to the same path as the sample solution and thus takes into account all parameters influencing the sample. © 1995 John Wiley&S

ISSN:0006-3592    

DOI:10.1002/bit.260450205

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractThe influence of the thermodynamic activity of water (aw)on the synthesis of triolein catalyzed byMucor mieheilipase was investigated. Its effect on the equilibrium and on the rates of the different reactions present, esteification and mono‐ and diglyceride isomerization, was evaluated through measurements made in controlled water activity atmosphere. The apparent equilibrium constants were measured from the concentration of the different species as a function of the intial glycerol‐to oleic‐acid ratio using all the values at once with a multi‐response nonlinear regression technique. Rate constants were determined from kinetic measurements and non‐linear regression uning the variation of the concentration of all significant species in the system. Except for the synthesis of diolein from monoolein, which shows a maximum foraw≈ 0.5, the apparent rate constants of the various reactions are not significantly affected by the value of the water activity. The equilibrium is shifted to‐ward the synthesis of triolein for low values ofaw, indicating that in the design of a process for triglyceride synthesis, usingM. mieheilipase as a catalyst, the water activity can be lowered to extreme values to favor the synthesis, without any sacrifice on the productivity of the process. © 1995 John W

ISSN:0006-3592    

DOI:10.1002/bit.260450206

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractA real‐time fuzzy‐knowledge‐based system for fault diagnosis and control of bioprocesses was constructed using the object‐oriented programming environment Small‐talk/V Mac. The basic system was implemented in a Macintosh Quadra 900 computer and built to function connected on line to the process computer. Fuzzy logic was employed in handling uncertainties both in the knowledge and in measurements. The fuzzy sets defined for the process variables could be changed on‐line according to process dynamics. Process knowledge was implemented in a graphical two‐level hierachical knowledge base. In on‐line process control the system first recognizes the current process phase on the basis of top‐level rules in the knowledge‐base. Then, according to the results of process diagnosis based on measurement data, the appropriate control strategy is subsequently inferred making use of the lower level rules describing the process during the phase in question. © 1995

ISSN:0006-3592    

DOI:10.1002/bit.260450207

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractInsulin or albumin was immobilized on collagen beads using water‐soluble carbodiimide. Adhesion of STO mouse fibroblast cells onto the beads decreased with increasing the amount of immobilized proteins. Growth of the cells was remarkably accelerated on the insulinimmobilized collagen beads, which can be used for serum‐free cell culture. The growth acceleration became larger with increasing the amount of immobilized insulin, while it became smaller with increasing the amount of immobilized albumin. In addition, the immobilized insulin more strongly accelerated the cell growth than free insulin plus collagen beads. © 1995 John Wiley&Sons,

ISSN:0006-3592    

DOI:10.1002/bit.260450208

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractThere are three major methods for buffer exchange of proteins at industrial scale: size exclusion chromatography (SEC), tangential flow filtration (TFF), and countercurrent dialysis (CCD). In order to determine the optimal technology for a given process, a study was done to compare these technologies on a technological and economic basis. This comparison required that new mathematical models be developed which enable the common features of each unit operation to be directly compared. The new concept of a diavolume equivalent for SEC, defined as the inverse of the fractional loading, was also introduced to aid in this comparison. Variables that were examined for each unit operation included range of buffer exchange, dilution of protein solution, yield, buffer requirements, total operating time, throughput, plant space, capital, raw materials, and labor costs. It was found that TFF and CCD have a greater range of buffer exchange than SEC. TFF also provides the advantage that concentration of the protein can readily be accomplished in the same step. For processes of equal batch size and yield, TFF and CCD also provide a two‐ to five‐ fold improvement in each of the remaining variables. The major economic advantage in using TFF and CCD over SEC is the decreased plant size required for manufacturing and thus the longer term use of existing facilities. Situations where SEC (or CCD) would be favored over TFF are when protein denaturation occurs in TFF but does not occur in SEC. © 1995 John Wiley&Sons,

ISSN:0006-3592    

DOI:10.1002/bit.260450209

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractLeukocyte filters are applied clinically to remove leukocytes from blood. In order to optimize leukocyte filters, a mathematical model to describe the leukocyte filtration process was developed by modification of a general theoretical model for depth filtration. The model presented here can be used to predict the time‐dependent leukocyte filtration as a function of cell–cell interaction in the filter, filter efficiency, filter capacity, filter dimensions, and leukocyte concentration in the suspension applied to the filter. The results of different leukocyte filtration experiments previously reported in the literature could be well described by the present model. © 1995 John Wiley&Sons,

ISSN:0006-3592    

DOI:10.1002/bit.260450210

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY

摘要:

AbstractWe report experimental data of mass transfer of oxygen into yeast resuspension in a pulsed baffled bioreactor. The bioreactor consists of a 50‐mm‐diameter column with the presence of a series of either wall (orifice) or central (disc) baffles or a mixture of both where fluid oscillation can also be supermposed during the experiments. Air bubbles are sparged into the bottom of the pulsed baffled bioreactor, and the kinetics of liquid oxygen concentration in the yeast solution is followed using a dissolved oxygen probe with a fast response time of 3 s together with the dynamic gassing‐out technique. Among the three different baffle geometries investigated, the orifice baffles gave the highest and sharpest increase in the oxygen transfer rate, and the trends in thekLameasurements are consistent with the fluid mechanics observed within both the systems and previous work. In addition, we have also compared thekLavalues with those obtained in a stirred tank; an 11% increase in theKLais reported. © 1995 John Wiley&Son

ISSN:0006-3592    

DOI:10.1002/bit.260450211

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1995

数据来源: WILEY