When working with small volumes evaporation of culture medium during incubation and disturbance of the atmosphere in the incubator can become a critical factor. Especially in 96-well plates with a working volume of 100-200 µL evaporation can lead to accumulation of media components, thereby affecting cell metabolism.

Filling these wells with medium without cells creates a more stable micro-environment in the plate and therefore minimizes the edge effect. In addition, opening the incubator while incubating the plates should be reduced as much as possible to not disturb the atmosphere inside.