Are mass spectrometry and sequencing possible after staining with GelCode Blue Stain Reagent?
Yes, see PNAS 98:12056-61 and J. Biol.Chem. 275:3462-8.
Can I re-use the stain for multiple gels?
For best results, use the stain solution only once. Lower amounts of staining may result when the stain solution is used more than once, producing variable results.
Can membranes be stained with GelCode Blue Stain Reagent?
Yes. The product instructions include a procedure for staining membranes. Wash membrane for 1-2 minutes in water, stain for 2-5 minutes and destain with 50% methanol, 1% acetic acid. For best results staining membranes, use MemCode Reversible Protein Stains for Membranes (Product # 24580, 24585).
Can a Western blot be performed on a gel that has been stained with GelCode Blue Stain Reagent?
Yes; however, transfer and subsequent probing efficiency may be decreased in some systems.
Is sensitivity of the GelCode Blue Stain the same for all types of proteins?
No. Some proteins can be detected to 8 ng. Most proteins are detectable at 25 ng. Some proteins (e.g., phosphoproteins) do not stain well.
Why are the phosphoproteins in my sample not staining well?
Coomassie dye does not bind well to phosphoproteins or other hydrophobic proteins. When using these proteins, switch to a silver stain to see total protein.
When I first started using this bottle of stain, it worked well, but now it is not working. What's wrong?
The bottle contents must be mixed well before dispensing to ensure a uniform solution.
Instead of performing the Water Wash Enhancement Step, may I destain with water/MeOH/acetic acid as I am used to doing with other coomassie stains?
Yes, any procedures that one ordinarily uses for coomassie-stained gels may be successfully performed on GelCode Blue Reagent-stained gels.
Can GelCode Blue Stain Reagent be used for isoelectric focusing (IEF) gels?
Yes. IEF gels can be stained, but they first must be fixed in 20% TCA for 30 minutes followed by washing with water for 30 minutes; then the gel can be stained as usual.
Can a GelCode Blue Reagent-stained gel also be stained with silver stain?
Yes. The GelCode Blue Reagent-stained gel must be fixed overnight with 30% methanol and 10% acetic acid, followed by washing with water for 1 hour; then the gel is then ready for staining with a silver stain (e.g., Product # 24612).
Do any gel types require fixing before staining?
Yes. For gels electrophoresed with MOPS or MES running buffers, fix gel with 50% methanol and 7% acetic acid for 15 minutes and extensively wash with water.
Is there a faster staining method using GelCode Blue Stain Reagent?
Yes. The product instructions include a microwave procedure for faster staining with only a minimal loss in sensitivity. Protein bands can be detected approximately 30 minutes after electrophoresis.
Does GelCode Blue Stain Reagent contain coomassie G-250 or R-250 dye and what is the difference?
GelCode Blue Stain Reagent contains coomassie G-250 dye. Coomassie G (green)-250 is greenish, stains bright blue, is less soluble and used in colloidal form (stain enters a protein better than the acrylamide, so background is minimized). Coomassie R (red)-250 dye is reddish, stains dark blue and is more soluble, usually resulting in high background.