SilverQuest™ Silver Stain Kit From Invitrogen

This is a product that Invitrogen markets for use over traditional methods of silver staining. Although Invitrogen states that you get similar results, I have found that traditional methods seem to be more sensitive in my hands. The major difference over the traditional method is that this kit does not use glutaraldehyde or formaldehyde (which modify lysine and arginine side-chain residues) in the sensitizer and as a result, the kit is safer. Also, and the biggest reason why I use this kit is that proteins stayed with this method of silver-staining can be directly analyzed via mass spectrometry. I have sent samples to our center for analysis using MALDI-TOF MS and have gotten really good results.

I have seen some silver-staining methods that take less than an hour and some protocols that take several hours. The SilverQuest™ instruction manual states that the staining can be completed in 90 minutes, but even after using this kit multiple times, it still takes me a little over two hours. Also, with traditional protocols you have to make the reagents fresh every time so you must take that into consideration. With this kit, all reagents are ready to go with minor additions and since you don’t have to work with glutaraldehyde or formaldehyde, you can pour the used reagents down the sink.

I have used this kit with protein samples that I have obtained from either immunoprecipitations or from direct protein pulldowns (GST-tagged protein as bait with cytosolic or nuclear protein preps). I strongly recommend that if you do a similar method that you WASH, WASH, WASH your IPS or pulldowns prior to running them on the gel. Your bait or antibody will be at a high concentration while what you are looking for will be relatively weak. When you start the developing portion of the protocol you will immediately see your bait or antibody and it will take longer for your unknown proteins of interest to appear. I have found that if you don’t wash enough, you will have a lot of ‘junk’ and high background show up on the gel.

The trouble-shooting section in the manual is definitely worth reading even before you run your gel. Also, the manual recommends for more sensitive results to leave in gel in fixative overnight. I really can’t tell if this is better or not. I traditionally have left my gels in fixative overnight anyway as I usually run the gel in the afternoon and don’t feel like staying around until 8pm at night. Also in the manual, there is a method for “fast” staining. I have not used this method so I can’t really say much about it.

I do not have any experience with testing this kit with “home-made” acrylamide gels. I have always used the NOVEX gels from Invitrogen with this kit. Invitrogen recommends using the NOVEX gels for mass spec analysis. I do feel that the NOVEX gels are of much higher quality than the “home-made” gels that I have used in the past. You might want to do a comparison for yourself.

Overall, this is a good kit. I strongly recommend it if you are planning on sending samples off for mass spec analysis.