Back to Basics CO2 Incubator: A Laboratory Staple

Research in the areas of cell biology, molecular biology, cancer, pharmaceuticals, etc. has made amazing strides during the past decade and the technology used in those areas has had to keep pace. The landscape (or labscape) of a typical life science laboratory has changed dramatically over the years but the CO2 incubator continues to be a staple in the research lab. Although the ultimate goal of maintaining cell culture stocks has not changed, the functioning and operation of CO2 incubators has become more accurate, more reliable and more convenient. CO2 incubators are designed with the user and his/her applications in mind. Three primary areas of concern when selecting a CO2 incubator are with reliability, contamination control, and ease of use. The following overviews several of the options that are available for CO2 incubators.

CO2 Control: The Options—Infrared or Thermal Conductivity

CO2 Incubator

CO2 levels within the chamber are established with a set point and are controlled to maintain that set point. When the door is opened CO2 escapes from the chamber and a CO2 sensor detects a drop in the level of CO2. CO2 is automatically injected to raise the level to the set point. Two control systems are available to detect changes in CO2 levels—Thermal Conductivity (TC) and Infrared (IR).

The thermal conductivity system works by measuring the resistance between two thermistors, one exposed to the chamber environment and the other enclosed. The presence of CO2 in the chamber changes the resistance between the two thermistors. The level of CO2 is determined by measuring the resistance. A drawback of the thermal conductivity system is that changes in temperature and relative humidity can affect the accuracy of the sensor. Frequent door openings, which cause fluctuations in temperature and relative humidity in addition to CO2 levels, can affect the accuracy of the thermal conductivity sensor.

The infrared system was developed as an alternative to the TC sensor and is a more accurate controller of CO2 levels. The infrared system detects CO2 levels with an optical sensor. An air sample from the chamber is passed between an IR emitter (a light source) and the sensor. The sensor detects a reduction in the IR from the emitter as the CO2 in the air sample absorbs the IR. The amount of IR absorbed is relative to the levels of CO2 in the air sample. The IR sensor is not effected by variations in temperature and humidity so it is more accurate than the TC sensor especially following door openings.  However, the IR system is usually more expensive than the TC system.

Temperature Control: The Options—Water-jacketed or Radiant-walled

Maintaining a constant temperature in the incubator is critical to the health and growth of cultured cells. There are two primary heating options to consider when selecting a small to mid-sized CO2 incubator: water-jacketed and radiant-walled. Although both heating systems are accurate and reliable, they each have their advantages and disadvantages.

Water-jacketed incubators maintain temperature by surrounding the interior chamber with heated water in a separate compartment. The water is heated and circulates around the inner chamber via natural convection. The heat from the water radiates to the interior chamber maintaining a constant temperature inside. Water is a particularly effective insulator and the water-jacket system is considered a more reliable method of heating in case of a power outage. In the wake of a power failure, a water-jacketed incubator will hold a set temperature inside the chamber 4-5 times longer than a radiant-walled unit.

Radiant-walled incubators heat the interior chamber using heaters mounted in the surrounding cavity that radiate heat through to the inside chamber. A radiant-walled heating system allows for quicker recovery of temperature following door openings or changes in temperature settings. Radiant-walled heating systems are also more simplified for the user, not requiring filling, monitoring, and emptying water in the water jacket.

A fan may be mounted outside of the culturing area to help to circulate the air inside the chamber without disturbing cultures. This gentle circulation speeds recovery of internal temperature as well as CO2 and humidity levels following door openings.

Humidity: Protecting Cultures from Desiccation

Desiccation results in lost cultures. It is important to maintain adequate moisture inside the chamber to prevent the drying out of cultures. Large CO2 incubators may use steam-generators or atomizers to control relative humidity levels but most small to mid-sized incubators use humidity pans to produce humidity through evaporation. Humidity pans produce relative humidity levels between 95-98%. Some incubators have a humidity reservoir that holds water in a heated pan. This increases evaporation. A humidity reservoir may increase the relative humidity levels to 97-98% but this system is more complicated and problems may arise due to an increase in the number of components and these units may be prone to internal "sweating".

Contamination: Exterminating the Bugs!

Contamination is a major source of frustration in cell culturing. Manufacturers of CO2 incubators have found ways to help combat contamination. By reducing the areas or surfaces where microorganisms can grow and by incorporating autodecontamination cycles are ways that manufacturers help researchers prevent contamination.  Many manufacturers also offer HEPA filters in CO2 incubators to reduce contamination during the incubation cycle. Copper-lined chambers with copper shelves and fixtures also reduce fungal growth and other contaminants. Removable shelves and crevice-free interiors or coved corners in drawn internal chamber reduce the areas where contaminants can begin growing. Surface areas are also more accessible for the use of disinfectants. Some CO2 incubators also have autodecontamination cycles that decontaminate the internal chamber in between incubation cycles. Autodecontamination works by raising the internal temperature to +90oC for several hours thereby killing contaminating microorganisms. The autodecontamination cycle, used in conjunction with HEPA filters, greatly reduces contamination.

Ease of Use

A CO2 incubator should be easy to use and maintain. With the introduction of microprocessor controls and various accessories, the CO2 incubator is approaching a "set it and forget it" mode of operation. Features such as over-temperature thermostats and alarms, CO2 alarms, door opening alarms, password protection of settings, self-calibration, and autodecontamination cycles offer easy operation and security for the user. Other features such as stackable models help conserve valuable lab space. Some models require stacking kits sold as accessories while other units are stackable without kits or tools. Some models also offer exchangeable door swings to accommodate lab space and placement.

Selecting the size of incubator to meet your storage needs and space limitations contributes to the ease of use. CO2 incubators range in size from "personal" bench-top incubators (<40 liters) to large-capacity incubators (>700 liters). The mid-sized models (140-180 liters) offer the most options described above but the smaller and larger capacity incubators accommodate specific storage requirements. The broad range of sizes, options, and accessories for CO2 incubators are quite numerous and should meet any researcher's needs.