changes in protein profiles in bortezomib applied multiple myeloma ...

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STOCK REAGENT PREPARATION (cont.)
10% Ammonium Persulfade (w/v)
Dissolve 100 mg ammonium persulfate in 1 ml distilled water
Sample Buffer (SDS reducing buffer: 62.5 Mm Tris- HCl, pH 6.8, 20% Glycerol, 2%
SDS, 5% β- Mercaptoethanol)
Distilled water 3.0 ml
0.5 M Tris- HCl, pH 6.8 1.0 ml
Glycerol 1.6 ml
10% SDS 1.6 ml
β- Mercaptoethanol 0.4 ml
0.5% (w/v) bromophenol blue (in water) 0.4 ml
Total 8.0 ml
Dillute the sample at least 1:4 with sample buffer. Heat at 95°C for 4 minutes.
5x Running Buffer (1x= 25 mM Tris, 192 mM glycine, 0.1% SDS, pH 8.3)
Tris base 45.0 g
Glycine 216.0 g
SDS 15.0 g
Distilled water to 3 L. Do not adjust the pH with acid or base. Store at 4°C.
Warm to 37°C before use if precipitation occurs. Dillute 300 ml 5x stock with
1.2 L distilled water for one electrophoretic run.
To prepare 100 ml of 1X TGS buffer, 20 ml of 5X TGS buffer was diluted to a
final volume of 100 ml with ultra pure water.
B.6.2.2. Preparation of Overlay Agarose
SDS Electrophoresis Buffer
Tris Base 3.03 g
Glycine 14.40 g
SDS 1.0 g
were dissolved in 1000 ml of distilled water
Overlay Agarose
Agarose 0.50 g
Bromophenol Blue 2.0 mg
SDS Electrophoresis Buffer 100 ml
stored at 4°C
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