Ethylenediaminetetraacetic acid, Coomassie Brilliant Blue R-250
, sodium dodecyl sulfate, lauric acid, potassium hydroxide, and ammonium hydroxide were analytical grade and supplied by Sigma-Aldrich.
The gel was finally stained with 0.25% Coomassie brilliant blue R-250
stain (Amresco, USA) for 4 hours and then suitably destained for best visibility of the protein bands with several changes of destaining solution and stored in 7% glacial acetic acid solution for photography.
Protein concentration was determined using Coomassie Brilliant Blue R-250
, and bovine serum albumin was used as the standard .
Protein pattern was observed by staining a gel with coomassie brilliant blue R-250
The plate was then stained for 24 hours using a solution containing 37% formaldehyde, 15 ml; methanol, 27 ml; water, 63 ml; and Coomassie brilliant blue R-250
, 2 mg.
The purified protein samples were seen as two or three smeared bands alter Coomassie brilliant blue R-250
staining (Figure 3A).
After electrophoresis, gels were stained with Coomassie brilliant blue R-250
. For determination of molecular weights, the gel was calibrated with the following marker proteins: 116, 97.4, 66.2, 37.6, 28.5, and 18.4 kDa (Sigma Chemical Company, St.
After electrophoresis the gel was stained with 0.1 % w/v Coomassie Brilliant Blue R-250
(Sigma) in methanol:water:ethanoic acid (5:5:1) and then destained with a solution of methanol:water:ethanoic acid (1:17:2).
The gels were washed in a 2.5% Triton X-100 solution with shaking for 30 min and then incubated in 50 mL reaction buffer (40 mM Tris-HCl, pH 8.0; 10 mM Ca[Cl.sub.2], 0.01% NA[N.sub.3]) at 37[degrees]C for 12 hr before staining with 0.25% Coomassie brilliant blue R-250
in 50% methanol and 10% acetic acid for 1 hr.
Coomassie Brilliant Blue R-250
stain was used to visualize the proteins.
Gels were stained with either Coomassie Brilliant Blue R-250
(Dzandu et al., 1984) or silver (Wray et at., 1981).