RESUMEN
The purpose of this article is to review the current state of large-scale cell culture in terms of its applications, problems, and potential. Because of the commercial and proprietary nature of most large-scale cell culture processes, this review does not contain many detailed scientific results although an attempt is made to address some key issues and findings. Much of this summary deals with processes having an established, commercial track record but some attention is given to more recent innovations with interesting potential applications. Reference is made to plant cell culture but the main emphasis is on mammalian cells.
Asunto(s)
Células Cultivadas , Animales , Anticuerpos Monoclonales , Hibridomas , Interferones , Perfusión , Proteínas Recombinantes , Tecnología , Vacunas ViralesRESUMEN
We present our rationale for attempting to cover some exposed cranial implants by tissue transfers. We have tried this in 9 patients, and we have had success in 5.
Asunto(s)
Cráneo/cirugía , Cirugía Plástica , Resinas Acrílicas , Humanos , Trasplante de Piel , Tantalio , Trasplante Autólogo , Cicatrización de HeridasRESUMEN
Osteonecrosis of the skull produced by an electrical burn presented as a dramatic "cold" bone lesion on an MDP bone scan, despite normal skull radiographs. Four months later the skull radiograph showed marked bone resorption, and the three-phase bone scan confirmed healing and new osteoblastic activity. The pathophysiology of high-voltage injuries is outlined.
Asunto(s)
Quemaduras por Electricidad/diagnóstico por imagen , Cráneo/patología , Quemaduras por Electricidad/patología , Humanos , Persona de Mediana Edad , Radiografía , Cintigrafía , Cráneo/diagnóstico por imagenAsunto(s)
Labio Leporino/cirugía , Fisura del Paladar/cirugía , Cirugía Plástica , Adolescente , Adulto , Cicatriz/crecimiento & desarrollo , Dentaduras , Femenino , Humanos , Labio/crecimiento & desarrollo , Masculino , Anomalías de la Boca/cirugía , Rinoplastia , Ajuste Social , Factores Socioeconómicos , Habla/clasificaciónRESUMEN
The serum requirement for the human lymphoblastoid cell line MICH can be replaced by iron, methylcellulose and bovine serum albumin. Fatty acid free preparations of albumin are inactive.
Asunto(s)
Sangre , División Celular , Línea Celular , Medios de Cultivo , División Celular/efectos de los fármacos , Cobre/farmacología , Ácidos Grasos/farmacología , Humanos , Hierro/farmacología , Metilcelulosa/farmacología , Albúmina Sérica Bovina/farmacología , Zinc/farmacologíaRESUMEN
Mammalian cell suspension culture systems are being used increasingly in the biotechnology industry. This is due to their many advantages including simplicity and homogeneity of culture. Suspension systems are very adaptable (e.g., for microcarrier, microencapsulation, or other methods of culture). Their engineering is thoroughly understood and standardized at large scale, and automation and cleaning procedures are well established. Suspension systems offer the possibility of quick implementation of production protocols due to their ability to be scaled easily once the basic culture parameters are understood. The only main disadvantage of the suspension culture systems to date is their inapplicability for the production of human vaccines from either primary cell lines or from normal human diploid cell lines (Hayflick et al., 1987 and references therein). One of the great advantages of suspension culture is the opportunity it provides to study interactions of metabolic and production phenomena in chemostat or turbidostat steady-state systems. Furthermore, in suspension culture systems from which cell number and cell mass measurements are easy to obtain, rigorous and quantitative estimations of the effects of growth conditions or perturbations of metabolic homeostasis can be made. Such studies can speed up the development of optimal processes. With our increasing understanding of factors influencing expression in mammalian cells (Cohen and Levinson, 1988; Santoro et al., 1988) and the direct application of new methods in suspension culture (Rhodes and Birch, 1988), its usefulness and importance is likely to increase in the future. In this chapter, we have described some of the potential uses of the various suspension culture systems and have covered most of the established technology and literature. Due to the rapid developments and needs in the biotechnology industry and the versatility of suspension culture systems, it is probable that many more variations on this theme will evolve in the near future at both the pilot and production scales.
Asunto(s)
Técnicas de Cultivo/métodos , Biosíntesis de Proteínas , Animales , Anticuerpos Monoclonales/biosíntesis , Anticuerpos Monoclonales/aislamiento & purificación , Biotecnología/instrumentación , Biotecnología/métodos , Línea Celular , Células Cultivadas , Técnicas de Cultivo/instrumentación , Fermentación , Humanos , Mamíferos , Proteínas/aislamiento & purificación , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/aislamiento & purificación , Vacunas/biosíntesis , Vacunas/aislamiento & purificaciónRESUMEN
Glucose limited the maximum population density of the cell line Bri 8 over a wide range of pH values. Efficiency of glucose utilisation varied with pH and hence pH influenced maximum population density. Surprisingly the ratio of glucose consumed to lactic acid produced was not greatly influenced by pH.
Asunto(s)
División Celular , Glucosa/metabolismo , Línea Celular , Medios de Cultivo , Humanos , Concentración de Iones de Hidrógeno , Cetoácidos/metabolismo , Lactatos/metabolismoRESUMEN
High speed continuous electrophoretic fractionation of crude commercial trypsin preparations has resulted in the resolution of the cell monolayer dispersal activity into three fractions, two of which were considered useful in the production of fully dispersed single cell suspensions. This separation could be achieved at an input rate for crude enzyme of 500 mg per minute.
Asunto(s)
Separación Celular , Tripsina/aislamiento & purificación , Línea Celular , Quimotripsina/metabolismo , Electroforesis , Humanos , Elastasa Pancreática/metabolismo , Tripsina/metabolismo , Tripsina/farmacologíaRESUMEN
Aggressive management of severe burns minimizes contractures and helps to maintain muscle tone, joint function and psychological well-being. The positioning, activity and exercise programs, splinting and bandaging, and skin care of burned children carried out by the burns team at the Hospital for Sick Children, Toronto is outlined.
Asunto(s)
Quemaduras/complicaciones , Contractura/prevención & control , Terapia por Ejercicio , Postura , Férulas (Fijadores) , Quemaduras/cirugía , Niño , Contractura/etiología , Humanos , Inmovilización , Trasplante de Piel , Trasplante AutólogoRESUMEN
Swine serum is an inexpensive substitute for foetal calf serum for lymphocyte culture. It gives an increased yield of Ig from 1788 cells and permits the large scale production of cells for Ig mRNA isolation.
Asunto(s)
Sangre , Inmunoglobulinas/biosíntesis , Linfocitos/citología , Animales , Bovinos , División Celular , Línea Celular , Medios de Cultivo , Sangre Fetal , Humanos , Linfocitos/metabolismo , ARN Mensajero/biosíntesis , PorcinosRESUMEN
Mouse hybridoma cells producing a monoclonal antibody to anti-B blood group antigen have been grown in airlift fermenters up to 30 litres capacity. As a prerequisite for scaling the process up, the oxygen utilization rate of the cells has been established by a dynamic method using an oxygen electrode. The data is compared with that obtained from a chemostat under oxygen limited steady state conditions.
Asunto(s)
Células Productoras de Anticuerpos/metabolismo , Hibridomas/metabolismo , Consumo de Oxígeno , Animales , Células Productoras de Anticuerpos/citología , División Celular , Línea Celular , Hibridomas/citología , RatonesRESUMEN
We have developed several approaches to create cell lines with improved characteristics in cell culture. In some cases it has been possible to isolate natural variants with useful properties. Cholesterol independent variants of the mouse NSO myeloma cell line were isolated by cloning in a selective medium. A glutamine independent variant of a hyridoma was isolated by continuous (chemostat) culture under glutamine limited conditions in the presence of glutamate. Choline independent cells were isolated from a choline limited chemostat. In an alternative approach to modifying cell behaviour, we have used recombinant DNA techniques to introduce the glutamine synthetase (GS) gene to a hybridoma. This resulted in glutamine independence and increased productivity.
Asunto(s)
Técnicas de Cultivo/métodos , Glutamato-Amoníaco Ligasa/biosíntesis , Proteínas Recombinantes/biosíntesis , Transfección , Animales , División Celular/efectos de los fármacos , Colesterol/farmacología , Colina/farmacología , Células Clonales , ADN Recombinante/metabolismo , Ácido Glutámico/farmacología , Glutamina/farmacología , Hibridomas/citología , Ratones , Mieloma Múltiple , Células Tumorales CultivadasRESUMEN
From recently published data on the amino-terminal structures of human and mouse interferons, we have predicted and synthesised an oligonucleotide capable of priming specifically the reverse transcription of human fibroblast interferon mRNA present within a total mRNA population. From these transcripts we determined the sequence of the 5'-terminus of the mRNA and identified a putative pre-peptide signal sequence. This enabled us to predict the sequence of another primer capable of directing the synthesis of interferon double-stranded cDNA corresponding to the entire coding region of the mRNA. Further sequencing studies also enabled us to establish the identity of 47 consecutive amino acids beginning with the methionine residue at the amino-terminus of the mature protein.
Asunto(s)
Fibroblastos/metabolismo , Genes MHC Clase II , Interferones/genética , Secuencia de Aminoácidos , Humanos , Oligodesoxirribonucleótidos , Oligonucleótidos , Poli T , ARN Mensajero/genética , ADN Polimerasa Dirigida por ARNRESUMEN
We have employed an inverse engineering strategy based on quantitative proteome analysis to identify changes in intracellular protein abundance that correlate with increased specific recombinant monoclonal antibody production (qMab) by engineered murine myeloma (NS0) cells. Four homogeneous NS0 cell lines differing in qMab were isolated from a pool of primary transfectants. The proteome of each stably transfected cell line was analyzed at mid-exponential growth phase by two-dimensional gel electrophoresis (2D-PAGE) and individual protein spot volume data derived from digitized gel images were compared statistically. To identify changes in protein abundance associated with qMab datasets were screened for proteins that exhibited either a linear correlation with cell line qMab or a conserved change in abundance specific only to the cell line with highest qMab. Several proteins with altered abundance were identified by mass spectrometry. Proteins exhibiting a significant increase in abundance with increasing qMab included molecular chaperones known to interact directly with nascent immunoglobulins during their folding and assembly (e.g., BiP, endoplasmin, protein disulfide isomerase). 2D-PAGE analysis showed that in all cell lines Mab light chain was more abundant than heavy chain, indicating that this is a likely prerequisite for efficient Mab production. In summary, these data reveal both the adaptive responses and molecular mechanisms enabling mammalian cells in culture to achieve high-level recombinant monoclonal antibody production.