How to properly sterilize a bioreactor?

The sterilization operation of the bioreactor is an essential link before carrying out the biological reaction experiment. If the sterilization operation is not in place, it is easy to contaminate the fermentation products during the operation of the bioreactor, resulting in greater losses. Therefore, it is incredibly important find the skills of sterilization,then we will briefly discuss the steps of sterilizing the bioreactor.
 

STEP 1.

Remove the stirring motor of the bioreactor, lay flat on the table, wrap the motor interface with tinfoil and bind it with a rubber band to prevent it from falling off during the sterilization process, so as to eliminate the damage to the motor interface caused by high-temperature steam generated during high-pressure steam sterilization.
 

STEP 2.

Remove the temperature sensor from the vessel. The temperature sensor does not directly touch the fermentation broth and does not require sterilization.
 

STEP 3.

To install the pH electrode:
  • There is a rubber ring protection between the pH electrode and the panel.
  • Be sure to tighten the upper and lower fixing nuts of the electrode.
  • The pH electrode cable interface is wrapped with tin foil and tied with rubber bands to prevent the sterilization process from falling off and affecting the performance of the electrode.
  • The pH electrode should be calibrated before sterilization. 
 

STEP 4.

To install the DO electrode:
  • There is a rubber ring protection between the DO electrode and the panel.
  • Be sure to tighten the upper and lower fixing nuts of the electrode.
  • Wrap the DO electrode cable interface with tin foil and tie it with a rubber band to prevent it from falling off during the sterilization process and affecting the performance of the electrode.
  • Calibrate DO electrode after sterilization.
  • Before sterilization, pay attention to whether the electrolyte in the dissolved oxygen electrode still exists, if not, add it.
     

STEP 5.

Remove the defoaming or liquid level cable, and insert the defoaming or liquid level cable to the bottom. After sterilization, it is positioned to the required height and then do not press it again. The height of the defoaming electrode is about 1.5cm from the liquid surface.
 

STEP 6.

Remove the cooling water inlet and outlet pipes on the bioreactor and the inlet and outlet pipes on the condenser in turn. When removing, clamp the joints of the removed water pipes with clips to prevent water from flowing out.
 

STEP 7.

Remove the air tube from the luer connector, and clamp the silicone tube of the air inlet with a hose clamp. The silicone tube and the air filtration are sterilized together with the glass vessel. The exhaust port is also clamped with a hose clamp, and then the silicone tube is sterilized with the air filtration. Put the exhaust filter into the sterilizer together.
 

STEP 8.

The mouth of the four-way feeding tube can be blocked with a silicone tube, or it can be connected to the feeding bottle for sterilization (one end of the feeding bottle is connected to the breathing valve), and it is processed according to the working conditions (if it is sterilized together with the feeding bottle, the feeding end must be sterilized together with the feeding bottle. Clamp with hose clamps).
 

STEP 9.

Use hose clamps to clamp the panel end of the harvest tube and the spare nozzle respectively.
 

STEP 10.

Before sterilization, the position of the stirring blade should be adjusted. The lower stirring blade should be installed at the bottom of the stirring shaft as much as possible, and the upper stirring blade should be adjusted to the middle position of the added fermentation liquid.
 

STEP 11.

Before sterilization, tighten the six retaining nuts of the bioreactor.
 

STEP 12.

  • Before sterilization, the flame inoculation port must be loosened to prevent the pressure difference between the inside and outside of the vessel from being too large.
  • Usually high temperature and high pressure sterilization at 121℃, 20min (the specific time is adjusted according to the medium composition).
  • After sterilization, the temperature of the fermenter may be high, do not touch it directly, wait for the equipment to cool down, and wear protective gloves before operating.
  • After sterilization, tighten the flame inoculation port immediately to prevent contamination.
     
The above content is the specific steps of sterilizing the bioreactor. Emphasize again: the step of sterilizing the bioreactor before using it is crucial to prevent microbial contamination when using the bioreactor.

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