Removal of animal cells and cell debris by continuous two-stage centrifugation

Abstract

The development of a continuous two-stage centrifugation process for the removal of hybridoma cells and cell debris from culture broth is reported. The aim was to generate supernatant, which can directly be used for ultrafiltration or chromatography without any filtration steps.

The first centrifugation step for the removal of hybridoma cells from repeated batch fermentation in 100 L scale was performed by an improved disc stack separator. The centrifuge was equipped with a hydrohermetic feed system, which allowed a damage free separation of the cells by moderate centrifugal force.

The clarification for mammalian cells in this step was in the range of 99 to 99.9 %. The amount of subcellular particles in the clarified liquid phase was reduced up to 50 % of the particle content in the feed stream.

In a second centrifugation step the removal of the remaining amount of cell debris and other small particles was carried out. It was performed with centrifugal forces up to 18000 • g in a high speed centrifuge equipped with a continuous flow system.

The monoclonal antibodies in the resulting supernatant were directly concentrated by ultrafiltration and Chromatographic methods. In this process no fouling of the UF-membranes or Chromatographic membranes was observed.

Costs for membrane replacement can be drastically reduced by employing the two-stage centrifugation process.