by Catherine Shaffer
In theory, a CO2 incubator is simply a heated box with a controlled environment and high humidity in which to incubate culture plates. And yet, in practice, an incubator is quite technologically complex. Manufacturers are constantly tinkering with the features, from the user interface to the basic heating and sensing functions. The problem, ultimately, with an incubator is that reality constantly intrudes on the platonically perfect environment inside. Each time the door opens, heat is lost and microbes are invited inside. Contamination and desiccation are frequent problems in incubators, and solutions for one often work in opposition to the other. For example, increased airflow for HEPA filtration helps with contamination, but can have a desiccating effect on the cell culture. As a result, some of the most sophisticated technology in the laboratory can be found in the humble incubator.
Choices in incubators
CO2 incubators can be broadly grouped into two categories: the water-jacketed and air-jacketed models. Traditionally, water-jacketed models have been preferred because they have maintained a more stable temperature. One major disadvantage of the water jacket, however, is that the temperature recovers slowly after the door is opened. Air jacketed models, particularly the newest technology, approach the temperature stability of a water jacket incubator, but recover quickly after the door is opened. An air jacket incubator is also faster and easier to set up, since it doesn't require an initial overnight equilibration step.
Although it would seem that air jacket is the wave of the future, this is not true in every case. Water jacket incubators still have some key advantages, and geography can be a surprising contributor to the choice. Says Buckner Richerson, vice president of international sales for NuAire, Inc., "The first thing I would say is it's just a matter of taste.... Both are excellent. From a global perspective...if you're in a country where ambient temp is very high, such as India, an unairconditioned lab can be 40°C, and the incubator is at 37, this is best handled by water jacket. It won't be affected as fast by ambient conditions as an air jacket." It is also easier to install cooling coils in a water jacket incubator than an air jacket incubator. On the other hand, Richerson cites Japan as a location where air jacketed incubators are preferred, since it is customary there to turn off or turn down the heat at night. A water jacketed incubator would not be able to react to these temperature fluctuations as effectively.
The water jacketed incubator still has a slight edge over the best air jacketed incubators, so for extremely sensitive applications, such as the incubation of human embryos for in vitro fertilization, many laboratories will choose a water jacketed incubator. For most applications, however, the choice comes down to preference.
Overcoming contamination
Contamination of cultures is a serious concern, especially when working with labor-intensive tissue culture experiments, where a contamination event can ruin weeks of work. Contamination can come from mycoplasmas, bacteria, molds, spores, yeast, or fungi, and can suddenly fill the whole incubator, or lurk invisibly in a crevice or a water pan. Popular anticontamination measures include heat sterilization, UV sterilization, and HEPA filtration. Decontamination can be a separate process from incubation, as in a heat sterilization cycle, or it can be a continuous process that occurs simultaneously with incubation, as with HEPA filtration. The use of copper in the lining and fixtures of the incubator discourages the growth of microbes as well.
NuAire's incubators are based on the company's successful line of biosafety cabinets, which use HEPA filtration to control contamination. Although it would seem ideal to carry out the entire incubation inside an actual biosafety cabinet, the air flow in such a cabinet leads to desiccation. Ideally, an incubator should be maintained at 95% relative humidity. NuAire incorporated the same HEPA filtration technology in its incubators, along with humidity controls designed to maintain the proper humidity level for cell growth. Says Richerson, "We simply used what works well in biosafety cabinets. The incubator is built like a biosafety cabinet. We employed HEPA filtration at the heart of the system. A constantly clean interior is the secret to eliminating cross contamination. Culprits are filtered away, and don't have a chance to cause damage." NuAire has been monitoring contamination events in its incubators, and has only had 70 incidents since 1986.
Sanyo is taking a novel approach to incubator decontamination systems. They are introducing an innovative technology with their new line of incubators (Sterisonic™ GxP)—hydrogen peroxide sterilization. Sanyo's current line of incubators uses a patented UV sterilization system. This is part of a strategy to minimize downtime for decontamination cycles. A heat decontamination process can take the equipment out of commission for up to twenty-four hours, and the high temperatures put a lot of wear and tear on the interior hardware, including the heat sensor, microprocessors, and filters. With H2O2 sterilization, the incubator is misted with hydrogen peroxide. Following the hydrogen peroxide treatment, the UV system is activated, causing the H2O2to revert to sterile water and trace oxygen, and completing the sterilization process. Says Deepak M. Mistry, strategic development & marketing manager for Sanyo, "We have a very unique solution to [decontamination]. But I guess what it really comes down to in this market is people are looking for a very reliable chamber that gives them the best, most stable and optimum culturing environment.... I think as a manufacturer, we're kind of given the burden to explain to them why our technology is better in that respect."
All the bells and whistles
In addition to robust heating and decontamination systems, researchers are also enjoying a range of new and improved features. Temperature and CO2 sensors are increasingly interfaced with alarm and backup systems. NuAire's newest line of incubators can detect when the CO2 sensor has gone out of range and activate a backup system to maintain the flow of CO2. Other features enhance usability, such as a touch screen interface on NuAire's newest line (due in 2010).
The incubation market is increasingly catering to more specialized and time-sensitive applications. Laboratories are asking for more performance from their incubators, larger chambers, more accurate and precise controls, more convenience, and less downtime. The CO2 incubator is an old laboratory standby, but it's worth looking at the newest generation, if only to save yourself the nightmare of yet another contamination crisis.