Sometimes less is more

The liquid volume for cultivation of respiratory active microorganisms like   Escherichia coli   should not exceed 5 – 10 % of the total flask volume (i. e. 10 to maximum 25 mL in a 250 mL flask) otherwise there may be a lack of oxygen supply.

Recommended culture conditions for the aerobic cultivation of  E. coli  in a 250 mL wide neck flask with cotton plug or membrane cap are:

• 10 mL filling volume 
  
• 300 rpm shaking speed 
  
• 50 mm shaking diameter
  

Microorganisms with minor breathing activity, like yeasts or filamentous fungi should be cultivated as follows:

• 250 mL wide neck flask with cotton plug or membrane cap 
  
• 25 mL filling volume 
  
• 300 rpm shaking speed 
  
• 50 mm shaking diameter
  

For plant or animal cell culture the following conditions should be chosen:

disposable shake flask with membrane cap or narrow neck flask with cotton plug
controlled humidity to reduce evaporation loss

• 50 - 100 mL filling volume
  
• 100 - 150 rpm shaking speed
  
• 50 mm shaking diameter
  

Sampling reduces the filling volume of the flask and thereby changes the cultivation conditions. Usually the shaker is stopped during the sampling procedure resulting in an interruption of oxygen supply. This may influence the total metabolic activity of the cells.

To minimize the impact of this procedure on experimental results an appropriate number of flasks should be scheduled. One entire flask can then be harvested for sampling, the shaker is stopped for only a short period of time and the oxygen supply of the cells is guaranteed.