Cells were then scraped into lysis buffer and lysed on ice for 20 min before conducting quantitative analysis as described above.
After washing with PBS, the cells were then scraped with lysis buffer containing 20 mM Tris-HCl (pH 7.5), 150 mM NaCl, 1 mM Na2EDTA, 1 mM EGTA, 1% Triton X, 2.5 mM sodium pyrophosphate, 1 mM β-glycerophosphate, 1 mM Na3VO4, and 1 μg/mL leupeptin.
Cells were then scraped into the RIPA buffer and homogenized by repeated pipetting.
The cells were then scraped and resuspended in lysis buffer (1% Triton X-100, 150 mM NaCl, 25 mM HEPES, 10 mM MgCl2, 1 mM EDTA and 2% glycerol).
The cells were then scraped and lysed in NP-40 lysis buffer (Sigma-Aldrich) and Western Blotting was done with appropriate primary and secondary antibodies as previously described (Rajput et al, 2013).
The cultured cells were then scraped and lysed in 100 μL of lysis buffer (20 mM Tris-HCl, pH 7.4, 0.05% Triton-X 100, and 2 mM EDTA).
The cells were then scraped and lysed in Nonidet P-40 lysis buffer.
The cells were then scraped in 200 ul ice-cold Cell Lytic M lysis buffer from Sigma and sonicated for 20 seconds, followed by centrifugation at 5000 g for 2 minutes at 4°C.
Cells were then scraped in ice-cold PBS, centrifuged and cell pellets lysed with 1% NP40 lysis buffer (10 m M HEPES pH 7.5, 0.15 m M NaCl, 10% glycerol, protease inhibitors cocktail and 1 m M PMSF).
Cells were then scraped, centrifuged at 600 g for 5 min, and resuspended in 650 μl PXL lysis buffer (PBS without Ca2+ and Mg2+, 0.1% SDS, 0.5% NP-40 and 1 50 RNase-free RQ1 DNase, and 1 100 RNase OUT).
