After the purified complexes were washed three times with lysis buffer, they were solubilized in Laemmli buffer, followed by boiling for 5 min at 100°C.
mda-7/IL-24 IBs were washed using '3 + 1' washing buffers (about 1 g IBs were solubilized in 30 mL washing buffer): buffer I, buffer II, buffer III, and deionized water (Table 2).
The washed inclusion bodies were solubilized in solubilization buffer (8 M urea, 200 mM β-mercaptoethanol, 20 mM Tris HCl, 3 mM EDTA, pH 8.0).
Then, the inclusion bodies containing the Cry1Ac protoxin were solubilized in solubilization buffer (10 mM DTT, 50 mM sodium carbonate, and pH 10.5) for 2 h, at 37°C, and dialyzed against water in a 12 kDa dialysis membrane.
For blue native electrophoresis, 100 µg mitochondria were solubilized in solubilization buffer: 1% (w/v) digitonin (Calbiochem), 20 mM Tris, pH 7.4, 0.1 mM EDTA, 50 mM NaCl, 10% (v/v) glycerol.
The beads were washed with 1% NP40 cell lysis buffer and bound proteins were solubilized in 2× NuPage LDS reducing sample buffer, boiled and loaded on 4 12% Bis-Tris Nupage gels (Invitrogen).
After TCA-precipitation pellets were solubilized in buffer A containing 1% SDS (20 min at 37°C), and diluted 10 times with buffer A (containing 0.2% Triton X100).
One milligram of protein sample, 7 μL of DTT (1 mol · L−1), and 1.75 μL of IPG buffer (20 mmol · L−1) were solubilized in 350 μL of rehydration solution containing 8 mol · L−1 urea, 2% CHAPS, and a trace of bromophenol blue.
Pellets bearing 70S ribosomes were solubilized in buffer A at 4°C and saved at −80°C.
Briefly, samples were solubilized in buffer containing Tris-HCl (pH 6.8), SDS, bromophenol blue, glycerol and 2-β-mercaptoethanol.
