Preparation of Extracts

To immunoprecipitate proteins from embryonic tissue or cell culture, the material has to be disrupted under conditions that solubilize the proteins (see Notes 3 and 4).

1. Embryos or cell cultures are collected and washed in PBS prior to homogeniza-tion using a dounce homogenizer, or other commercial disruption device, with ice-cold RIPA or NP40 lysis buffer.

2. Centrifuge the lysate at 12,000g for 10 min at 4°C to remove the insoluble material; in the case of NP-40 lysates, this will also cause the unlysed nuclei to sediment.

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