Cell Culture FAQ: What impact does your CO2 incubator have on reproducibility?

In biological systems, high reproducibility of results is much harder to achieve than in physical or chemical experiments. Several factors contribute to the variation between individual results:

• The quality and age of cells
• The sterility of the cultures
• The consistency and quality of equipment and media used
• The individual person performing the experiment
• The duration of the experiment

In addition to these factors, the incubator plays a crucial role as it provides the ideally uniform culture conditions needed for optimal and standardized cell growth. The homeostasis inside the incubator provides and influences three factors of the cell environment:

• A precise temperature control
• A CO2 atmosphere that regulates the pH in bicarbonate-buffered culture media
• A humid atmosphere to limit evaporation from the culture vessels

This homeostasis, however, is disturbed every time the inner incubator door is opened as the air inside the incubator flows out. Afterwards, all three factors need to recover to the set specifications which might take several minutes. In order to counteract the unavoidable effects of opening the doors, you should consider the following:

• Avoid opening any door for longer than 30s
• Shorten the number of times the door is opened as much as possible
• Limit the angle the door is opened
• Look for an incubator with fast recovery rates after door opening
• Use an incubator with divided doors which greatly reduces the disturbance and saves gas consumption
• Consider a second incubator when the number of times the door is opens exceeds the acceptable amount
• Consider a small additional incubator for sensitive long-term experiments
• Consider making a tally sheet to count daily door openings