Oxygen consumption profiles of Sf-9 insect cells and their culture at low temperature to circumvent oxygen starvation

Abstract

The oxygen consumption profile of Sf-9 insect cells was determined under various conditions using a 250 ml spinner flask. The specific oxygen consumption rate, q, of Sf-9 was scarcely changed with cell density and growth phases except for stationary phase. However, the q was varied with dissolved oxygen (DO) concentration and described by a Monod-form equation in the range of 20–34 °C. For uninfected Sf-9, the maximum specific oxygen consumption rate, q max, in the equation decreased with decreasing temperature, whereas the saturation constant, K S, remained constant independent of temperature up to 28 °C, at which Sf-9 cultures were usually performed. This indicated that the cellular physiology of Sf-9 was not altered in the temperature region. On a viral infection, the q max increased by 30–40% irrespective of temperature, and the K S increased more largely at higher temperature. The virus-infected Sf-9 culture was performed at 20 and 28 °C using a 2 l spinner flask with a working volume of 1 l, and DO behavior and recombinant protein production were determined. The oxygenation was done by supplying only air to mimic a large-scale culture of oxygenation-limited conditions. As a result, only the 20 °C culture was successful to keep DO concentration above the critical DO concentration (3 K S) and produce a large amount of green fluorescent protein, a recombinant model protein, but not the 28 °C culture. The DO behavior in both the 20 and 28 °C cultures was well simulated by model calculation using the oxygen consumption parameters obtained above. These results indicated that a low temperature culture was useful to circumvent oxygen starvation of virus-infected Sf-9 cells and to successfully produce recombinant proteins.