It is pipetted gently on top of the culture medium, where it forms a continuous layer because the silicone oil is less dense than aqueous medium and does not mix with it. The layer needs to cover the whole liquid surface to be effective, so dispense slowly and check coverage before starting a long acquisition.

Silicone oil is permeable to oxygen and carbon dioxide, so gas exchange continues, though more slowly than through an open surface. For CO2-buffered media in long experiments this is usually acceptable, but pairing the oil with a stage top incubator or a HEPES-buffered medium gives more predictable pH control.

Yes. The oil is optically clear and, in the inverted microscope configuration used with ibidi labware, the light path goes through the chamber bottom rather than the oil layer in any case. It does not degrade image quality.

No. It is a biocompatible silicone oil supplied sterile and validated for direct contact with cell culture, and it is compatible with all ibidi products and standard culture applications.

Whenever the surface-to-volume ratio is high or the experiment is long. Small-volume formats such as µ-Slides and 96-well µ-Plates lose a significant fraction of their medium over several days, and edge wells in multi-well plates are affected first. If you have seen unexplained drift in cell behaviour late in a time-lapse, evaporation is a likely culprit.