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Separating Gel Buffer or Lower Tris

• 1.5 M Tris-HCl pH 8.8
• 0.4% SDS

Stacking Gel Buffer or Upper Tris

• 0.5 M Tris-Cl, pH 6.8
• 0.4% SDS

Acrylamide Stock

• 30% (w/v) acrylamide
• 0.8% (w/v) bis

Sample Buffer- 2X

• 50% Stacking gel buffer
• 20% glycerol (v/v)
• 4% (w/v) DTT or 10% (v/v) ß-mercaptethanol
• 0.02% Bromophenol blue

Running Buffer
For 1x - 0.025 M Tris-glycine, pH 8.9
0.1% SDS
For 5x - 0.125 M Tris-glycine, pH 8.9
0.5% SDS

Coomassie Stain
To make Coomassie staining solution you will need:

• 400mL of water
• 500mL of methanol
• 1g of Coomassie Blue
• 100mL of acetic acid

1. Mix together for one hour
2. Add 100mL of acetic acid
3. Filter solution into another container
4. Stain gel(s) for 1 hour

Destaining Protocol
To make destaining solution you will need:

• 600mL of water
• 300mL of methanol
• 100mL of acetic acid
• Kim wipes, or some other absorbent material

Destain for 1 hour, occasionally removing/ replacing kimwipes as they become saturated w/ Coomassie.

In-Gel Digestion Protocol
Short and long methods of in-gel digestion

TBS (Tris Buffered Saline)

• 10 mM Tris pH 7.4
• 150 mM NaCl

TBS-Triton

• 10 mM Tris, pH 7.4
• 150 mM NaCl
• 0.05% Triton X-100

Blocking Buffer

• 10 mM Tris, pH 7.4
• 150 mM NaCl
• <3% to 4% BSA (Bovine Serum Albumin) or 5 &endash; 8% milk./li>

Ponceau S Stain

• Make 1: 10 dilution in water from concentrate
• Ponceau S 2%
• Trichloroacetic acid 30%
• Sulfosalicylic acid 30%