Incubation

1. Take eggs from the cool store, and leave on bench to reach room temperature before setting in the incubator.

2. When embryos are being collected and fixed directly, or used to provide material for culture, the eggs can be incubated vertically. Lay eggs on their sides when incubating for manipulations (see Chapters 16-19 and 22), since during incubation, the yolk will rotate such that its least dense region, the embryonic blastoderm, rotates to the highest point: the upper side of the egg, which is where a window will later be cut through the shell. This property of rotation is retained in older embryos.

3. For best viability, incubate eggs at high relative humidity (>50%) and in the range of 37.5-39°C. Times to reach a specific developmental stage depend on the exact temperature (which will vary with position within the incubator) and duration of previous storage (old eggs have an appreciable lag-phase before they resume development). If older embryos (more than 9 d of incubation) are required, rocking of the eggs during incubation improves viability by preventing adhesion between extraembryonic membranes and the shell (egg incubators are commercially available for this purpose). However, rocking does not appreciably increase viability of younger embryos, and these can be incubated in conventional ovens or incubators that function with high internal humidity.

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