Siver Stain Protocol

(from Dunbar, B.S. 1987. Two-Dimesional Electrophoresis and Immunological Techniques. Plenum Press, NY. 372 pp)

Solutions

Solution A:

• 0.8 g silver nitrate
• Bring to 4 mL in distilled water

Solution B:

• 21 mL 0.36% NaOH
• 1.4 mL 14.8 M ammonium hydroxide
• For best results, make fresh daily

Solution C:

• add Solution A dropwise to B with constant stirring then bring to 100 mL with distilled water.
• NOTE: SOLUTION C MUST BE USED WITHIN 5 MIN

Solution D:

• 2.5 mL 1% citric acid
• 0.25 mL 37% formaldehyde
• Make to 500 mL with distilled water.
• SOLUTION D MUST BE “VERY FRESH”

Misc Solutions:

• #1: 50 % methanol
• #2: 5% acetic acid

Protocol

1. Soak gel in 50% MeOH at least 60 min
2. Stain gel in Solution C (15 min with gentle agitation)
3. Wash gel in water (5 min with gentle agitation)
4. Develop gel in Solution D
5. Stop with 5% acetic acid
6. Wash gel with water and place in 50% MeOH overnight.