Author: Jacob Peter Matson; Amy M. House; Gavin D. Grant; Huaitong Wu; Joanna Perez; Jeanette Gowen Cook
Title: Intrinsic checkpoint deficiency during cell cycle re-entry from quiescence Document date: 2019_2_22
ID: dsbucda9_41
Snippet: To prepare total protein lysate for immunoblot, cells were harvested with trypsin and frozen in dry ice, then lysed in cold cytoskeletal buffer (CSK) (10 mM Pipes pH 7.0, 300 mM sucrose, 100 mM NaCl, 3 mM MgCl2 hexahydrate) with 0.5% triton x-100 (Sigma Aldrich) and protease and phosphatase inhibitors (0.1 mM Pefabloc, 1 µg/mL pepstatin A, 1 µg/mL leupeptin, 1 µg/mL aprotinin, 10 µg/mL phosvitin, 1 mM β-glycerol phosphate, 1 mM sodium orthov.....
Document: To prepare total protein lysate for immunoblot, cells were harvested with trypsin and frozen in dry ice, then lysed in cold cytoskeletal buffer (CSK) (10 mM Pipes pH 7.0, 300 mM sucrose, 100 mM NaCl, 3 mM MgCl2 hexahydrate) with 0.5% triton x-100 (Sigma Aldrich) and protease and phosphatase inhibitors (0.1 mM Pefabloc, 1 µg/mL pepstatin A, 1 µg/mL leupeptin, 1 µg/mL aprotinin, 10 µg/mL phosvitin, 1 mM β-glycerol phosphate, 1 mM sodium orthovanadate) on ice for 15 minutes. Lysate was centrifuged at 13,000 x g at 4°C for 10 minutes, and a Bradford Assay (Biorad) was done on the supernatant to load equal amounts of protein per sample.
Search related documents:
Co phrase search for related documents- dry ice and phosphatase inhibitor protease: 1
- equal amount and protein lysate: 1
- β glycerol and sample protein: 1
- phosphatase inhibitor and sample protein: 1
- phosphatase inhibitor protease and sample protein: 1
Co phrase search for related documents, hyperlinks ordered by date