Since the trypsin digested peptides are very small in size, Is it possible to run them in SDS-PAGE and stain them (Silver stain).? Have anyone ever tried this method?
Hi Boopathi, you can run 25% + SDS page gels to isolate peptides. I would not recommend Ag staining because it makes the peptides insoluble.
For small peptides, the Tris-Tricine SDS-PAGE method of Schaegger and von Jagow is a good choice.
http://www.sciencedirect.com/science/article/pii/0003269787905872
Hi Boopathi, are your peptides larger than 1kDa? Adam's reference is good for those peptides. Coomassie staining should work.
Dear Steingrimur,
My peptides are less than 1kDa. Initially, I will perform the proteolysis by trypsin and i will go SDS-PAGE for the digested peptides.
I have managed to visualize peptides as small as 2kda and maybe slightly smaller but less than 1 kDa is extremely difficult. Also be careful about your choice of staining method as some colloidal coomaissie stains are not mass spec compatible. Why do you want to run the peptides on a gel ? is it for purification or separation purposes ?
Hi Gerry,
I'm trying to visualize the trypsin digested proteins in SDS-PAGE during control and treated conditions. In order to see the changes in banding patterns and also to purify the peptides.
For such small peptides, you should not use SDS-PAGE. You should use reverse-phase HPLC (C18 column, water-acetonitrile gradient with 0.1% trifluoroacetic acid, 214 nm UV absorbance detection).
- Badges
- Science method