VWR Life Science RIPA Lysis Buffer, Biotechnology Grade

Protein isolation Mammalian cells - Caco-2

Experiment
Protein isolation Mammalian cells - Caco-2
Product
VWR Life Science RIPA Lysis Buffer, Biotechnology Grade from VWR
Manufacturer
VWR

Protocol tips

Protocol tips
After cell harvesting, the pellets were washed three times with cold PBS, lysed with RIPA buffer containing protease inhibitors, and sonicated using a sonicator

Publication protocol

After cell harvesting, the pellets were washed three times with cold PBS, lysed with RIPA buffer (Ameresco, USA) containing protease inhibitors (Ameresco, USA), and sonicated using a sonicator. Protein concentrations were measured using a Pierce BCA protein assay kit (Thermo Scientific, USA). The cell lysates were subjected to polyacrylamide gel electrophoresis (PAGE, resolving gel 12% and stacking gel 4%). Then, the partitioned molecules were transferred to PVDF membrane by electrical blotting. The membranes were incubated with Odyssey blocking buffer for 1 h at room temperature, and then incubated with appropriate primary antibodies at 4oC overnight. After that, the membranes were incubated with accordant secondary antibodies at room temperature for 1 h. Band visualization and quantitation were carried out using a LI-COR Odyssey Imager (Li-COR, USA).

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