Zinc Negative Stain

This procedure is rapid and sensitive, able to detect 5 - 10 ng protein per band. The proteins are not themselves stained but show as clear transparent bands on an otherwise solid white gel background.

1. After SdsPage rinse the gel in distilled water for 30 seconds.
   
2. Incubate the gel in 0.2 M imidazole, 0.1% SDS for 15 minutes.
   
3. Discard the previous solution and incubate the gel in 0.2 M zinc sulphate until the gel background becomes a deep white consistency, about 30 seconds
   
   • protein bands will be transparent and colourless
     
4. Stop the stain by rinsing with copious distilled water.
   
   • be careful not to overstain the gel
     
5. Take a picture of the gel by placing it on a dark background exposed to white light.
   

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Notes

• Proteins are immobilized in the stain if the gel is kept hydrated. Stable for years.
  
• If dried, the stain will fade. Rehydration will restore the stain.
  
• Stained gels do not transfer well in semi-dry blotting apparatus. In submerged transfer apparatus, stained gels will transfer well if a zinc chelator like glycine is added eg: 25 mM Tris-HCl, 192 mM glycine pH 8.3 transfer buffer.
  

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References

Castellanos-Serra L, Proenza W, Huerta V, Moritz RL, Simpson RJ. Electrophoresis 20:732-737 (1999)

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