Overview
• Generate distinct nuclear and cytoplasmic fractions from cultured cells in 90 minutes
• For adherent or non-adherent cultured cells
• Compatible with downstream Western blotting applications
Detailed Description
This kit isolates proteins from both the cytoplasmic and nuclear fractions of cultured cells. The cytoplasm and the nucleus are separated from one another by simple centrifugation steps following lysis in a non-ionic detergent. The entire procedure can be performed in less than 1.5 hours. Kit includes Fractionation Buffer 1, 1 ml, Fractionation Buffer 2, 1.5 ml, Fractionation Buffer 3, 30 ml, and Nonidet® P-40 Substitute, 0.1 ml. Contains sufficient material for 20 extractions of 1 x 106 cells.
Resuspend cell pellet in a cytoplasmic lysis buffer for 15 minutes. Add detergent and pellet the nuclei. Transfer the supernatant containing the cytoplasmic fraction and further isolate the proteins with cytoplasmic extraction buffer. After the nuclei have been washed, the pellet is resuspended in nuclei lysis buffer for 15 minutes. The nuclei debris is pelleted and the supernatant contains the nuclei protein fraction.
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