Simultaneous cell disruption and aqueous two-phase extraction for isolation of intracellular recombinant proteins

A new technique was developed for the integrated processing of cell disrupt ion and aqueous two-phase extraction in a high-speed bead mill to separate intracellular proteins from microbial cells. The process was named as simul taneous cell disruption and aqueous two-phase extraction (SDATE). Advantage s, such as high cell disruption efficiency, biochemical activities preserva tion of proteins, cell debris elimination, and preliminary purification of the target protein were being claimed. When this technique was employed for isolating recombinant Tumor Necrosis Factor (TNF) from E. coli, overall pr otein concentration and TNF activity were found to have been increased. Mor e than 95% of TNF was partitioned into the top phase and all cell debris we re in the bottom phase. The partition coefficient was greater than 3 and th e TNF purification factor was greater than 6. It is shown that less separat ion steps were being utilized in the new technique, meaning a reduction in separation time and less process extractors required.