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Temperature control review for cell biology imaging

Temperature controller for biological experiments on microscope

We review here the existing equipments to perform temperature changes for the cell biology microscope imaging

Temperature homogeneity

A strong difference between the temperature displayed by a temperature controller and the effective temperature of your sample can occur in temperature control systems. Particularly when using immersion objectives, temperature shift as high as 10°C can be observed with most temperature control systems. When using DIC illumination, temperature shift up to 18°C have been locally observed in less than 1 minute [1]. Those temperature gradients of several degrees in the observation field can have a strong influence on the biological sample behavior.

[1] O. Buchner, C. Lütz and A. Holzinger, Journal of Microscopy 225, 183 (2007)

Speed of the temperature change and temperature range

Depending on the temperature control system used, the time required to perform a temperature change can vary from 30 minutes to 5 seconds. Basic temperature controllers for cell biology imaging using only joule heating elements enable temperature change from ambient to 100°C. For experiment requiring a temperature below the ambient, one should prefer systems using Peltier devices which generally enable temperature from -20°C to 100°C.

Compatibility with high resolution microscopy

Oil or water immersion objectives are commonly used for high resolution imaging in cell biology. They act as a heat sink for the biological sample since there are in direct contact with the glass coverslip. With some temperature controllers, temperature shifts higher than 10°C can appear in the imaging area where the objective enters in direct contact with the sample. These temperature shifts are always a percentage of the temperature difference between the sample temperature and the temperature of the objective. One approach to limit this problem is the use of additional temperature controlled objective collars which thermalize the body of the objective. Besides homogeneity problems, the thermal expansion of the sample cell should be minimized to keep the focus during high resolution microscopy. When using immersion objectives, the closer is the temperature control from the sample, the better is the homogeneity on the sample.

Multi temperature screening and control experiment

The investigation of temperature control effect on biological functions requires to get several temperatures on the sample at the same time to get reliable control experiments. This capability to submit the same sample to different temperature controls at the same time in identical conditions is necessary to ensure that the change in the biological system is due to temperature.

Artifacts

Some temperature controllers induce strong thermal artifacts and require knowledge of the physical phenomenon for a right interpretation of biological results. The use of immersion objectives or DIC illumination can generate strong temperature shifts on the sample depending on the chosen temperature controller. Classic joule heating or Peltier based systems can also experience overshooting problems, especially when the temperature control sensor is placed far away from the sample.

Biological compliance of temperature control systems

Temperature controllers can stress cells by mechanical deformation or flow shear stress. For example, some temperature controllers using thermalized micro-perfusion generate a strong flow rate directly on cells, creating shear stress, flushing of endocrine/paracrine signaling and being useless with non-adherent cells. On the other hand, using micro-perfusion enables you to provide gas and nutriments to the cells, which can increase the long term viability of the biological system under investigation. Depending on the temperature control, some biological systems will not be compatible.