A new wireless system for decentralised measurement of physiological parameters from shake flasks

Abstract

Background: Shake flasks are widely used because of their low price and simple handling. Many researcher are, however, not aware of the physiological consequences of oxygen limitation and substrate overflow metabolism that occur in shake flasks. Availability of a wireless measuring system brings the possibilities for quality control and design of cultivation conditions.

Results: Here we present a new wireless solution for the measurement of pH and oxygen from shake flasks with standard sensors, which allows data transmission over a distance of more than 100 metres in laboratory environments. This new system was applied to monitoring of cultivation conditions in shake flasks. The at-time monitoring of the growth conditions became possible by simple means. Here we demonstrate that with typical protocols E. coli shake flask cultures run into severe oxygen limitation and the medium is strongly acidified. Additionally the strength of the new system is demonstrated by continuous monitoring of the oxygen level in methanol-fed Pichia pastoris shake flask cultures, which allows the optimisation of substrate feeding for preventing starvation or methanol overfeed. 40 % higher cell density was obtained by preventing starvation phases which occur in standard shake flask protocols by adding methanol when the respiration activity decreased in the cultures.

Conclusion: The here introduced wireless system can read parallel sensor data over long distances from shake flasks that are under vigorous shaking in cultivation rooms or closed incubators. The presented technology allows centralised monitoring of decentralised targets. It is useful for the monitoring of pH and dissolved oxygen in shake flask cultures. It is not limited to standard sensors, but can be easily adopted to new types of sensors and measurement places (e.g., new sensor points in large-scale bioreactors).

Figure 1

SENBIT system. A) operation principle B) SENBIT device (receiver, shake flask system and transmitter).

Figure 2

pH and dissolved oxygen tension in a typical E. coli shake flask cultivation measured with SENBIT^®. The short periods of oxygen decrease are caused by stops of the shaker due to sampling. Strain RV308 was cultivated in mineral salt medium (baffled 1 l shake flask with 200 ml medium, 37°C, 180 rpm shaking, antifoam 0.1 ml l^-1).

Figure 3

Optimisation of substrate feeding in Pichia pastoris cultures. Pichia uses methanol as a carbon/energy source. Methanol, however, is toxic at high concentrations. A recombinant strain of Pichia pastoris GS115 was cultivated in 1 l baffled shake flasks with three side necks containing 200 ml of BMM medium at 30°C with 200 rpm. To maintain the expression of the product, 100 % of methanol was added to a final concentration of 0.5 % (v/v) either twice a day (a) or alternatively, at the time when the DOT level increased (b). Times for substrate feeding are indicated with vertical lines.