CPMSC Facility: Protocols and Forms
Mass Spectrometry Analysis Form
Proteomics Sample Submission Form
These procedures are compatible with mass spectrometry analysis of proteins. Quantities are for large (20 cm x 20 cm) gels.
Vorum Silver Stain Method
1. Fix 2-18 hrs. (50% Methanol/12% Acetic acid, 0.05% formalin), 1 liter/gel
2. Wash three times in 35% Ethanol/water for 20 min
3. Wash twice in distilled water for 10 minutes
4. Sensitize 2 min with 0.5L of 100 mM sodium thiosulfate, 30 mM potassium ferricyanide
5. Wash in 0.5 L water for 10 min. four times
6. Stain 20 min. with 1 L 0.2% silver nitrate, 0.076% formalin
7. Wash in water 1 min. twice
8. Develop till dark enough with 1 L of 6% sodium carbonate, 0.05% formalin, 0.0004% sodium thiosulfate
9. Stop 5 min. with 1 L of 50% Methanol/12% Acetic acid
10. Store in 1% Acetic acid/water
Desiderio Silver Destain
1. Water 5 min (3X)
2. Destain whole gel 15 min. with 1:1 v/v 30mM potassium ferricyanide and 100 mM sodium thiosulfate prepared fresh
3. Water 5 min. (10X)
4. Proceed to fixation procedure for staining or store at 1% Acetic acid/water
*Destain is useful if background darkens faster than the protein bands or if the gel is not stopped quickly enough and is generally too dark.
1. Vorum and Blum procedures: Presented at 48th American Society for Mass Spectrometry Conference on Mass Spectrometry June 11-15, 2000, Long Beach, CA, poster TPE 191. Now described in Lin JF, Chen QX, Tian HY, Gao X, Yu ML, Xu GJ, Zhao FK. Anal Bioanal Chem. 2008 Apr;390(7):1765-73 in a comparison of stains.
2. Desiderio procedure: R.R. Becklin, E.S. Umstor, D.M. Desiderio, poster WPF 216, 48th American Society for Mass Spectrometry Conference on Mass Spectrometry June 11-15, 2000, Long Beach, CA.
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