Silver Staining Procedure
来自 : 蚂蚁淘
Make up following solutions: (maybe stored for months at room temp.) Ratios are v/v.
Fixative: | 40% methanol; 10% acetic acid; 50% water |
Wash Sol.: | 30% ethanol in water |
Water: | d.i. water |
Reductant: | 200 mg sodium thiosulfate in 1 liter water |
Silver stain: | 2 g silver nitrate; 200 uL formaldehyde in 1 liter of water. Wrap bottle in aluminum foil to prevent light from getting in. |
Developer: | 30 g sodium carbonate; 5 mg sodium thiosulfate; 500 uL formaldehyde in 1 L water |
Stop Sol.: | 5% acetic acid in water |
Begin staining (Procedure is for mini-gel; takes about 2 hours; use longer incubation times for thicker gels)
- Put your gel in a large glass tray with about 1 liter volume capacity; use enough of each solution at each step to cover gel; agitate gently during steps; only touch gel with a glass rod or not at all until staining is finished; use large amounts of water for each washing step to give clean results!
- Fix proteins in gel using fixative for 20 minutes (gel can be left in here overnight etc?
- Wash three times in Wash Sol. (about 20 minutes each)
- Reduce in Reductant for 1 minute
- Wash three times in water (about 30 seconds each)
- Stain with Silver Stain for 20 minutes
- Wash three times in water (about 30 seconds each)
- Develop in Developer until bands become visible
- Wash three times in water (about 30 seconds each)
- Stop in Stop Sol. (Bands may fade if left in this solution; dry gels or save in sealed plastic bags at four degrees)
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