This Nuclear/Cytosol Extraction Kit provides a complete system that enables the separation of nuclear extract from the cytoplasmic fraction of mammalian cells. The optimized reagents and procedures provided with the kit allow separation of nuclear and cytoplasmic fractions quickly with little or no cross-contaminations. The extracted nuclear and cytoplasmic protein fractions are functional and compatible with downstream assays such as transcriptional activity, RNA splicing, gel shift assay, reporter assays, enzyme activity assays, and Western blotting.