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In vivo-like Cell Culture Conditions

For most cell types, it is necessary to work with incubator conditions directly on the microscope. The ibidi product line of Heating & Incubation Systems allows for the control of temperature, CO2 and O2 concentration, and relative humidity of live cell samples on the microscope. With our fast and precise regulations of all incubation parameters we can provide stable conditions for cell culture directly on the microscope.

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Temperature Homogeneity and Stability

The metabolism of cells is influenced by temperature. Therefore, observed parameters like prolifera­tion, adhesion, protein expression, and others will be temperature-dependent. Results observed at different temperatures are not considered to be comparable findings. All live cell imaging assays, even short-term assays are recommended to be performed at optimal cell conditions.

ibidi’s product line of Heating & Incubation Systems provides a stable and homogenous temperature in the incubation unit on the microscope. Temperature stability is not only important for cell culture conditions, but also very important in maintaining focus stability. The ibidi Heating Systems control temperature in an analog manner with a permanent, smooth regulation.

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concentration.jpgCO2 Concentration

The CO2 concentration in human exhaled air is 4 % to 5 %, which indicates that the CO2 dissolved in the blood is in equilibrium with air containing this CO2 concentration. Bicarbonate buffered culture media are normally designed to exhibit a pH of 7.4, at a CO2 concentration of 5 % in ambient air.

When using bicarbonate buffered media, the CO2 concentration in the ambient air of 5% is a com­mon standard in cell cultures. For these (and hypoxia) conditions, the ibidi Gas Incubation System will upgrade the ibidi Heating System to a complete stage top incubator.

Humidity and Evaporation

Salt and protein concentrations are crucial to natural and reproducible cell behavior. Evaporation increases the concentrations of substances in cell cultures in an undefined way, especially in low-volume cultures.

To minimize evaporation, the relative humidity of ambient air should be controlled to nearly 100%. The average values reached in cell culture incubators are between 80 - 95%. All ibidi Gas Incubation Systems provide a patented feed­back-controlled humidity regulation.

The design of the μ-Slides and μ-Dishes is optimized for low evaporation. For long-term studies, evaporation can additionally be reduced through the use of silicone oil, e.g., ibidi’s Anti-Evaporation Oil. It is overlaid onto the culture medium, and thus decreases evaporation. Silicone oil is highly gas permeable for O2 and CO2 but it blocks humidity loss very effectively. It is non-toxic and has been tested for biocompatibility with different cell types, including HT-1080, MDCK, NIH3T3, and HUVEC.

The in vivo O2 concentration in most types of tissue is around 2-5 kPa, which corresponds to 2-5 % oxygen in air. Therefore, cell culture experiments at oxygen levels of 21 % are highly artificial.

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CO2 and O2 Control

When combined with the ibidi Gas Incubation System, the ibidi Heating Systems are fully functional, stage top incubators with full CO2 and humidity control.
The CO2 / O2 version of the ibidi Gas Incubation System allows you to control the oxygen in the cell culture. This function is especially beneficial, because most cells cultured in vitro live at a much lower oxygen concentration in vivo.

ibidi Heating
Systems

ibidi Gas
Incubation System

ibidi OPAL – Optical O2 Measurement System

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The in vivo O2 concentration in most types of tissue is
around 2-5 kPa, which corresponds to 2-5 % oxygen in
air. Therefore, cell culture experiments at oxygen
levels of 21 % are highly artificial.

O2 Monitoring and Intracellular Measurement

In addition to controlling the oxygen concentration in the stage top incubator, it is indispensable to know the real oxygen concentration near the cells (monitoring extracellular O2), or even inside the cells (intracellular O2). Because cells consume oxygen, the concentrations are typically much lower in cell clusters, such as tissue or spheroids.

With the ibidi OPAL – Optical O2 Measurement System, you can measure the real oxygen concentration directly inside the Petri dish.

NOTE: The O2 concentrations are gas phase concentrations, and are specified in vol. %. This is not an accurate unit for O2 dissolved in water and should be converted to mg/l or µmol/l, using solubility tables. However, to keep things simple, we say “the liquid contains 20% of oxygen”, which actually means “the liquid is in equilibrium with ambient air, which contains 20 vol % of oxygen”.

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The extra- or intracellular oxygen concentration
is measured by
beads (left) or by a cell permeable
O2-sensitive fluorophore (right).