Valuation of growth parameters in monolayer keratinocyte cultures based on a two-dimensional cell placement model.

The influence of inoculum size on the growth of keratinocyte cells was investigated in a monolayer culture with serum-free medium. A growth model of cell placement was applied to the expression of the cell adhesion phase after the inoculation, lag phase, exponential growth phase, and stationary phase because of contact inhibition at high cell densities. Based on the model, the lag time until the onset of cell division was shortened in proportion to the logarithm of the inoculum cell size, resulting in the enhancement of overall cell propagation.

On-line monitoring of adhesion and proliferation of cultured hepatoma cells using optical waveguide lightmode spectroscopy (OWLS).

Monitoring of cell adhesion, cell spreading, and cell proliferation opens attractive perspectives in the on-line control of monolayer cell cultures in toxicity tests, in bioreactors as used for the serial production of skin grafts, or in extracorporeal liver devices. In this study the hepatoma Hep G2 cell adhesion and proliferation was monitored using an integrated optical method, optical waveguide lightmode spectroscopy (OWLS). This method is based upon refractive index measurements within a 100-nm thin layer above a Si(Ti)O(2) surface on which the cells were cultured and exposed to cytotoxic and cytostatic agents.

Optical waveguide lightmode spectroscopy (OWLS) to monitor cell proliferation quantitatively.

The use of microscopic observations used for in situ monitoring of cell proliferation in the production of epidermal autografts is not satisfactory. In particular, the identification of the projected cell area from microscopic pictures by image analysis (IA) depends on intensity edges and level of contrasts and is thus limited to subconfluent cultures. Some of these problems can be solved by using optical waveguide lightmode spectroscopy (OWLS), which measures the effective refractive index of a thin layer above an Si(Ti)O(2) waveguide surface.

Optical waveguide lightmode spectroscopy as a new method to study adhesion of anchorage-dependent cells as an indicator of metabolic state.

Optical Waveguide Lightmode Spectroscopy (OWLS) is based on measurements of the effective refractive index of a thin layer above the waveguide. Its potential as a whole-cell biosensor was demonstrated recently monitoring adhesion and spreading of Baby Hamster Kidney (BHK) cells on-line. In this work the OWLS is shown to be a promising tool to study the adhesion, morphology and metabolic state of fibroblasts in real time.

Development of an on-line monitoring system of human keratinocyte growth by image analysis and its application to bioreactor culture.

Human keratinocytes were cultured in serum-free medium for the purpose of on-line cell growth monitoring by image analysis. The validity of a process using a newly developed video microscopy system with image analysis for growth-rate monitoring in real time was verified by the measurement of the degree of confluence of keratinocytes in T-flasks and Petriperm dishes. The growth rate of keratinocytes was calculated subsequently from the linear relationship between average degree of confluence and cell concentration.

Computer controlled bioreactor for large-scale production of cultured skin grafts.

KERATOR--an automated membrane bioreactor--was developed to produce Autologous Wound Dressing (AWD) at significantly reduced cost and time of transplantation down to two weeks time. At the same time, the risk of human error is largely eliminated. The computer-controlled reactor is modular, allowing the production of up to 0.

Automated production of cultured epidermal autografts and sub-confluent epidermal autografts in a computer controlled bioreactor.

The objective of this work was to engineer an automated system for the production of cultured epidermal autografts and sub-confluent cultured epidermal autografts. Human epidermal cells were grown directly on a transparent FEP film, which was held in place and surrounded by a polycarbonate growth chamber. The growth chambers were stacked to accommodate various surface area requirements.

A novel culturing and grafting system for the treatment of leg ulcers.

The purpose of this study was to develop and test an efficient culturing and grafting system for the treatment of leg ulcers. The culturing system consisted of a Petriperm culture vessel (20 cm2) aseptically placed in a larger standard Petri dish (60 cm2). Skin cultures were established and cultivated in the Petriperm dish.

Kinetics of Human and Bovine Serum Albumin Adsorption at Silica-Titania Surfaces.

The interaction of proteins with surfaces is important in separation and purification procedures as well as in metabolism and its regulation. The degree of binding to a given surface in principle depends on the precise amino acid composition of the protein, although very little is presently known about the relationship between amino acid sequence and binding. Here we report accurate measurements of the kinetics of adsorption of two closely homologous serum albumins (human and bovine) to a hydrated metal oxide surface, using an accurate integrated optics technique.

Optical method for measurement of number and shape of attached cells in real time.

A novel, on-line method of accurately parametrizing the shape of spreading cells is described. The substrate on which the cells are deposited serves as an optical waveguide. The method is based on the interaction between the evanescent part of the guided waves and the cells.

Measurement of adhesion and spreading kinetics of baby hamster kidney and hybridoma cells using an integrated optical method.

A novel optical method was used for quantitative characterization and continuous measurement of both the adhesion and spreading of mammalian cells on inorganic surfaces. It is based upon the effective refractive index change caused by cells when they adhere to a planar optical waveguide. We have applied this technique to measure the kinetics of the adhesion and spreading processes of baby hamster kidney (BHK) cells adhering to surfaces coated with fibronectin and under different culture conditions (PBS, medium, serum, EDTA).

Kinetics of adhesion and spreading of animal cells.

The adhesion and spreading of cells at surfaces are well established phenomena which have been extensively observed using light or electron microscopy. The kinetics of both processes can be quantitatively measured by allowing the cells to attach themselves to the surface of a planar waveguide allows the number of cells per unit area and a parameter uniquely characterizing their shape, such as the area in contact with the surface, to be determined. Cells suspended in nutrient medium or pure phosphate buffer were allowed to attach to and spread on a metal oxide surface or a layer of fibronectin, and the kinetics of attachment and spreading have been measured.

A simulation model for the continuous production of acetoin and butanediol using Bacillus subtilis with integrated pervaporation separation.

The potential for producing acetoin and butanediol with a Bacillus subtilis strain was investigated with continuous culture using molasses as carbon substrate. The steady-state results were influenced by both oxygen and undetermined limiting compounds. Employing the known metabolic pathways, four overall stoichiometry relations were used with an energetic assumption on the energy requirements for biomass formation to establish a linear relations were used with an energetic assumption on the energy requirements for biomass formation to establish a linear relation between the overall rates, whose parameters were determined by linear regression.

Development of an enzyme membrane reactor for treatment of cyanide-containing wastewaters from the food industry.

Cyanidase, an immobilized enzyme preparation for hydrolyzing cyanide to ammonia and formate, was applied for the treatment of cyanide-containing waste waters from the food industry. Apricot seed extract was chosen as a model effluent. The enzymatic hydrolysis of pure amygdalin, the main cyanogenic glycoside in the extract, and the degradation of the cyanide formed was investigated and compared with the behavior of the real extract in a batch slurry reactor.

Kinetics of enzymatic degradation of cyanide.

Cyanidase is a new enzyme preparation capable of degrading cyanide in industrial wastewaters to ammonia and formate in an apparently one-step reaction, down to very low concentrations. This enzyme has both a high selectivity and affinity toward cyanide. A granular form of the biocatalyst was used in a recirculation fixed bed reactor in order to characterize the new biocatalyst with respect to pH, ionic strength, common ions normally present in wastewaters, mass transfer effects, and temperature.

Performance of membrane fixed biocatalyst reactors. I: Membrane reactor systems and modelling.

Enzymatic membrane reactors are discussed according to the state of biocatalyst and driving force of reaction. Particular attention is given to the Capillary Membrane Fixed Enzyme Reactor (CAMFER) for its favorable characteristics. It is shown that, for a practical range of operation conditions, both kinetic and mass transfer effects must be considered simultaneously.

Formation of oligosaccharides during enzymatic lactose hydrolysis and their importance in a whey hydrolysis process: Part II: Experimental.

Enzymatic lactose hydrolysis using two yeast and two fungal lactases that are of current technical interest was studied. The enzymes were compared regarding their oligosaccharide production. Parameters influencing oligosaccharide formation, together with the effect of immobilization were examined and conditions minimizing oligosaccharide content in the hydrolysis product were proposed.

Formation of oligosaccharides during enzymatic lactose: Part I: State of art.

Enzymatic lactose hydrolysis by beta-galactosidase (lactase) was investigated with respect to the formation of oligosaccharides. An analysis of the formation of oligosaccharides and their control is important in the development of technical applications for enzymatic lactose hydrolysis. The available literature data on transfer reactions of lactase were reviewed, compared, and presented in a concise tabular form.

Enzyme adsorption in porous supports: local thermodynamic equilibrium model.

Enzyme adsorption from a finite bath (batch adsorption) onto porous spherical supports is investigated both experimentally and theoretically using beta-galactosidase and Duolite ion-exchange resin as a model system. Efficient numerical techniques are presented that have been used in conjunction with a parameter estimation routine to evaluate adsorption isotherm constants. Results show that even for adsorption processes lasting almost 10 h, the majority of the enzyme is confined to the outer half of the support and, for high initial enzyme concentrations in the bath, this loading takes place as a slowly moving front.

Solubility and selective crystallization of lactose from solutions of its hydrolysis products glucose and galactose.

A high degree of conversion is desired when lactose is hydrolyzed to glucose and galactose. This produces, however, a high concentration of galactose, which is inhibitory for the enzyme catalyst (beta-galactosidase). The inhibition can be reduced by limiting the conversion per pass over the enzyme (e.