Best Practices: Measuring CO2, Humidity and Temperature in Life Science Incubators

Credit: Vaisala

by Jacalyn Whitney, Application Sales Engineer, Vaisala

Life science innovation depends on accurate incubator environment measurements. From the success of experiments to the viability of biological samples, incubators demand specific environmental conditions to facilitate optimal cell and microorganism growth and development — and viable outcomes.

Life science incubators provide carbon dioxide control in a humidified atmosphere with a constant temperature to maintain the most favorable environment for cell growth. Even the slightest fluctuations in CO₂, humidity and temperature can significantly impact sample viability and experimental results. Consequently, incubators must remain sealed properly to prevent outside contaminants from affecting the samples. And accurate incubator measurements can help ensure the success of your experiments, increase the reproducibility of your results, and promote the safety of researchers and staff.

In this article, let’s explore critical life science incubator measurement parameters, the importance of correctly installed, high-quality measurement solutions and what to look for when determining the right solution.

What is Measured in Laboratory Incubators?

Utilized across various life sciences applications, including cell culture, microbiology and molecular biology, laboratory incubators enable biological samples to grow and develop under specific conditions. The key parameters typically measured in laboratory incubators include temperature, humidity, and CO₂ concentration, but pressure and oxygen also play a part.

Temperature

Temperature is one of the essential parameters to measure in an incubator, as it directly affects the growth rate and viability of biological samples. Different types of incubators are designed to maintain specific temperature ranges, depending on the application. Inside a CO₂ incubator, for example, the air is heated and circulated to create a uniform temperature, with the average temperature being 98.6°F (37°C). On the other hand, refrigerated incubators used for microbiology often maintain a temperature range of 14° and 39° F.

Carefully controlled temperature can ensure even heating—and even moisture and CO₂ concentration in the air volume.

Humidity

Relative humidity also plays a critical role in applications where samples are sensitive to drying out. A high-humidity environment can help prevent evaporation and maintain the integrity of the samples. Most incubators utilize a water pan or atomizer used to humidify the cabinet. In an ideal case, the humidity inside the cabinet would be near 100% to keep the samples fresh and alive. However, relative humidity is often closer to 90% due to temperature gradients and the limited accuracy of moisture control. When water is slightly colder than the rest of the incubator, condensation can be avoided, but maintaining optimal humidity levels remains challenging — even with humidity-controlled atomizers generating water particles.

Complicating the issue, some sealing materials used for chamber wall or cable feedthrough sealing can cause humidity drift, increased response time and hysteresis of the humidity instrument. In chambers used for very dry conditions, some sealing materials absorb humidity and will act as a moisture buffer. Even relatively small amounts of these materials can delay the drying of the chamber.

CO₂ Concentration

Far and away the most demanding element of parameter control is CO₂ concentration. If the CO₂ concentration is too high, the pH of the media will become too acidic, which can damage the cells. If the concentration is too low, on the other hand, the media’s pH will become too basic, which can also cause harm to the cells.

Inside the incubator, pure 100% carbon dioxide is fed from a bottle and mixed with the cabinet air to achieve the required concentration. Most incubators require a 5% CO₂ concentration for cell culture, but a higher 10% concentration is occasionally used in some applications. CO₂ measurement and control capabilities typically range between 0% and 20%.

Pressure and Oxygen

The average pound per square inch over/under in an incubator is 15, while some incubators require oxygen to be calibrated and stabilized before CO₂ to ensure accurate CO₂ readings. Other parameters that may be measured in laboratory incubators include light intensity and shaking speed (in shaking incubators).

Most life science incubators are equipped with humidity and CO₂ sensors and controllers to maintain specific levels and a controlled environment. Accurate measurement and control of these parameters are critical to the success of experiments and the viability of biological samples.

The Value of Accurate CO₂ Measurement

Accurate CO₂ measurement is critical for the success of cell culture experiments and the reproducibility of experiments. The substances held in these chambers are extremely valuable and cannot risk tampering. Since samples are susceptible to environmental change, obtaining precise measurements within a small range is vital.

Challenges of Achieving Accurate CO₂ Measurements

Successful incubator measuring depends on several aspects, but achieving accurate measurements in incubators can be challenging due to various factors.

Incubators are hot, humid and require regular cleaning to avoid the growth of harmful microbes like bacteria, viruses, fungi and more. Different measurement sensors and technologies used in incubators can be damaged or destroyed by cleaning and sterilization methods, so operators must confirm with manufacturers before cleaning, wiping, sterilizizng or decontaminating. Plus, the space inside incubators needs to be optimized, so sensor limitations exist, but environmental parameters like temperature and pressure still must be compensated for.

Because gases like CO₂ are compressible, their mole densities change with changing ambient atmospheric pressure and temperature, meaning the output is temperature and pressure dependent. The partial pressure of each gas is the product of its volume concentration and the system's total pressure. Temperature compensation is also essential because the mole density increases as temperature decreases. That said, CO₂ measurement needs to be balanced if the measurement conditions deviate significantly from the calibration conditions.

Carbon dioxide measurements made at temperatures and pressures different from the calibration conditions may need to be corrected to achieve the required accuracy. The simplest form of correction for a volume percent reading can be done using a formula according to the ideal gas law:

c_corrected(%/ppm) = c_measured(%/ppm)*(1013*(t(°C)+273)) = (298K*p(hPa))

Another way to compensate for temperature is to connect special humidity and temperature probes—like the Vaisala HUMICAP Humidity and Temperature Probe HMP77B—alongside the CO₂ probe to automatically compensate for the CO₂ reading.

Another challenge is improper installation. High-quality instruments must be installed correctly for proper operation. Locate the CO₂ sensor in a representative, open place—away from the gas inlet. The sensor should be located away from hot and cold spots, but incubator corners may not suffice due to insufficient air circulation. Sealing the sensor thermally from the external environment (the outside areas of the chamber) is also vital to avoid humidity condensation and unwanted water.

Accurate, continuous and reliable measurements that detect any interference or change inside the incubator are vital in preventing costly consequences. But not all measurement solutions are created equal.

Best Practices to Ensure Accurate Incubator Measurements

Since changes in temperature or humidity can affect the accuracy of measurements inside the incubator, achieving accurate measurements in incubators requires careful attention to some best practices.

Choosing, and Calibrating, the Right Sensors

Long-term measurement performance can be achieved by selecting the proper measurement technology and regularly calibrating installed sensors. Remember that even the highest-quality instruments need regular maintenance and calibration. Over time, sensors that measure temperature, humidity, CO₂ concentration and other parameters can drift out of calibration, leading to inaccurate measurements. Validating the instrument’s measurements is also a best practice to ensure it performs according to expectations.

But not all measurement solutions are created equal. Instruments used to measure parameters inside incubators may have limitations that affect the accuracy of the measurements, such as sensors that may not be able to accurately measure critical parameters outside of a particular range.

Proper Sensor Placement and Installation

As noted, the location of sensors within the incubator affects the accuracy of measurements. Sensors should be placed in areas that are representative of the overall environment inside the incubator but not near hot and cold spots or gas inlets. Proper sensor installation also helps to ensure that the sensors are not affected by external factors, such as drafts or changes in ambient temperature.

Avoid Condensation When Sampling

Incubators and environmental chambers are challenging to measure, with gas samples usually drawn from environments with high temperatures and high humidity into room-temperature settings, which results in condensation. Whether using diffusion or a pump-aspirated sampling method, precaution must be taken when drawing gas samples from humid environments to protect the surfaces of the many sensors inside probes and the pumping chambers from condensation. Consider Nafion sample tubing as a solution that quickly moves water through the membrane wall and evaporates it into the surrounding air via pervaporation.

Accurate incubator measurements necessitate careful attention to sensor selection and installation, regular calibration, monitoring of environmental conditions and preventing condensation from damaging the CO₂ sensor. Researchers must understand the limitations of different instruments used to measure parameters inside the incubator and take steps to minimize human error and sample interference to drive reliable, reproducible results.

Unlocking CO₂ Measurement Excellence

Accurate measurement of parameters like CO₂ concentration, temperature and humidity propels the success of life science experiments in incubators. With deviations delivering false results and severe implications, proper installation and maintenance of high-quality measurement solutions help ensure accurate and reliable measurements that increase the reproducibility and overall success of experiments. Investing in high-quality CO₂ measurement solutions will not only provide better results but will also lead to increased confidence in research outcomes—ultimately contributing to incredible advancements in the field of life sciences.

About the author
Jacalyn Whitney is currently an Application Sales Engineer at Vaisala. She worked for Vacuum Barrier Corporation for more than three years prior to joining Vaisala. She holds a bachelor’s degree in mechanical engineering with a concentration in manufacturing and industrial design with a minor in mathematics from Merrimack College in Massachusetts.