Assuntos
Anticorpos Monoclonais/biossíntese , Reatores Biológicos , Hibridomas/imunologia , Animais , Anticorpos Monoclonais/efeitos dos fármacos , Substitutos Sanguíneos/farmacologia , Bovinos , Técnicas de Cultura de Células/métodos , Fluorocarbonos/farmacologia , Glucose/metabolismo , Hemoglobinas/farmacologia , Ácido Láctico/metabolismo , Oxigênio/farmacologia , Células Tumorais CultivadasRESUMO
Avid AL is an affinity chromatographic gel based on a synthetic ligand which has been designed for the purification of immunoglobulins. The binding capacities of the gel for human IgG were investigated under different conditions; various salts; salt concentrations; different buffer systems with pH ranging from 3.8 to 10.0; and, tissue culture supernatant supplemented with 20% serum. At a similar ionic strength, the binding capacity changed according to the salting out effect of the ions used. Kinetic parameters of the affinity adsorption showed that a strong salting out effect significantly decreased the dissociation constant. Avid AL gel purified IgG from human serum with a capacity of 26.2 mg/ml gel using binding buffer containing sodium sulfate. Recombinant Protein A gel was used as a control. In this study it also exhibited enhanced binding capacity with a high salt concentration binding buffer.
Assuntos
Cromatografia de Afinidade , Imunoglobulina G/metabolismo , Piridinas , Sefarose/análogos & derivados , Azidas , Humanos , Concentração de Íons de Hidrogênio , Imunoglobulina G/isolamento & purificação , Fenolsulfonaftaleína , Ligação Proteica , Sais , Azida SódicaRESUMO
Cell culture in hollow fiber bioreactors has been used as a method for large-scale production of monoclonal antibodies, viruses, cell-associated proteins and cancer antigens. We have examined an important variable in culturing cells in hollow fiber bioreactors: bioreactor composition. Eight different bioreactor designs which varied in nominal molecular weight cutoff, surface area, fiber material and ultra filtration rate were compared in large-scale hollow fiber cultures. A standard protocol utilizing the hybridoma 3C11 (ATCC HB 8511) or African green monkey kidney cells (Vero cells, ATCC CRL 1587) was designed so that the only variable examined was the hollow fiber bioreactor in use. The results suggest that surface area has little effect on antibody productivity, while fiber composition and ultrafiltration rate may play an important role. Vero cell growth was affected by both fiber composition and ultrafiltration rate.
