Biology reagent for cells, Extraction reagents for cells

Extraction Buffer for Mammalian Cells

This product is supplied as a ready-to-use solution for the lysis of the cultured mammalian cells. Proteins can be extracted by adding this buffer [R0246] to the cells and the extract can be used directly for further analysis such as western blotting. This product does not include protease inhibitors. Please add a protease inhibitor cocktail, if necessary.

Product

R0246 RIPA Buffer (Ready-to-use) [for Protein extraction]

Application

Add the following protease inhibitors to RIPA buffer [R0246].

Leupeptin 10 µg/mL
Pepstatin A 1 µg/mL
Aprotinin 3 µg/mL
AEBSF 1 mM
1. Wash the cultured mouse myeloma-derived cell sp2/0 twice with PBS.
2. Remove PBS and add 200 µL of either cold RIPA buffer [R0246] containing protease inhibitors or the other manufacturer's RIPA buffer containing the same protease inhibitors to 1.0 x 10⁶ cells.
3. Incubate the cells for 15 minutes on ice.
4. Centrifuge the cells at 10,000 x g for 10 minutes at 4 °C
5. Measure the protein concentration of the supernatants.
6. Analyze the supernatants using western blotting.

The extracts were transferred to a PVDF membrane after electrophoresis.
Anti-β actin antibody was used for detection.
Equal or better detection was observed than that of the other manufacturer's product.