CULTURE OF HYBRIDOMA IN DIALYSED MEDIA

ABSTRACT Hybridoma cells were cultivated in RPMI 1640 medium supplemented with a serum substitute, primatone and pluronic F68. After 52 hours of batch culture, the cells die rapidly. This observation was attributed to either the exhaustion of nutrients in the culture medium or to the release of toxic compounds by the cells. The “exhaustion” hypothesis was ruled out by supplementing the medium with glutamine, glucose and amino acids, but no improvement in cell viability at 52 h was observed. As regards toxicity, neither the addition of ammonium salts, of lactate at concentrations similar to those found in 52 h batch cultures decreased cell viability. On the contrary, dialysis of the 52 h and 72 h culture medium against complete medium (cut off 10 000 d) permetted normal growth. In an attempt to scale up this dialysis procedure, a hollow fiber dialysis system (ENKA) was connected to a 2 liters fermentor (Biolafitte). The described cultivation process allows to increase the concentration of living cells as well as concentration of immunogloblulins. Key words : Hybridoma, dialysis cartridge, conditioned medium.