Hydrodynamics inside wells of a 24-square deepwell microplate
For 24-square deepwell microtiter plates, culture volumes between 2.5 ml and 3.5 ml are generally used**, either at 300 rpm on a shaker with a 25 mm amplitude or at 225 rpm on a shaker with a 50 mm amplitude.
Under both conditions oxygen transfer rates for 2.5 ml cultures are approximately 35-39 mmol O2 l-1 h-1 (corresponding to k La values of 180-200 h-1).
** preferably use a culture volume of 2.5 ml, especially when the sandwich cover has to be take off occasionally (e.g. for sampling or feeding)
* Duetz et al. (2004) Biochemical Engineering Journal 17: 181-185 (data determined enzymatically using glucose oxidase, horse radish peroxidase, and ABTS)
The mixing times under the above conditions are less than a second, as determined by adding a drop of acid to an alkaline solution (containing a pH indicator), and following the colour change with a high-speed camera.
To obtain higher oxygen-transfer rates (50 mmol O2 l-1 h-1 or higher), shaking at 50 mm/300 rpm is possible. Preferably do not use higher cultures volume than 2.5 ml per well (2 ml may be even better) to prevent the culture fluid to touch upon the sandwich cover (and so possibly cause cross contamination when the cover is taken off)