Can I stain a gel with coomassie or other colorimetric stains after using this kit?
Yes. Fluorescence does not interfere with colorimetric staining methods.
Can a gel stained with the the 6xHis Staining Kit be used subsequently for Western blotting?
Do I need to adhere to the volumes listed in the instruction book?
These volumes are suggested based on a typical 8 x 10 cm gel. If you are using a smaller or larger gel, adjust the volumes accordingly.
Does the size of the expressed tagged protein make a difference?
No, it should not.
How can I make a record of the staining results?
A CCD camera works well. Typically, a two-second exposure with the aperture open is sufficient time.
How can I visualize my 6xHis-tagged proteins with this staining kit?
UV transilluminator light box wavelengths between 280-320 nm work well. Proteins appear as a fluorescent yellow when exposed to UV light.
How long does this staining procedure require?
Staining takes approximately two hours.
How sensitive is the 6xHis Tag Stain?
It will detect 0.2 µg of protein or more.
Is fixation necessary before staining?
Is this staining procedure compatible with tris-tricine gels?
My protein has been purified over a metal-ion affinity column. Can it be used directly?
We recommend dialyzing your sample before electrophoresis and staining. Slide-A-Lyzer Dialysis Cassettes, Slide-A-Lyzer MINI Dialysis Units and SnakeSkin Dialysis Tubing are excellent tools for dialysis.
What is the purpose of this product?
Fusion tags, such as a string of six histidines (6xHis), can facilitate the purification and detection of proteins of interest. Minimal addition of extra amino acids that are poorly immunogenic does not interfere with protein folding. The Pierce 6xHis Protein Tag Staining Kit is designed to detect 6xHis-labeled proteins in SDS-PAGE gels, eliminating the time necessary to blot and detect protein extracts with antibodies.
What is the thickness of gel that is suitable for staining?
Gels that are 0.75 mm to 1.5 mm thick have been shown to allow adequate signal resolution.
Why don't I see staining of my 6xHis protein or protein control?
Because SDS interferes with staining, it is important that you wash the gel well with deionized water before staining. The minimum detection limit is 0.2 µg of a 6xHis-labeled protein. Most commercially available markers contain 6xHis proteins in such low concentration that they cannot be detected with this kit. For best results, use the 6xHis Protein Control Set (Product # 24572, included in the kit 24575).
Will it also stain 3xHis-, 4xHis-, 5xHis- and 10xHis-tagged proteins?
Based on the chemistry, we believe that it may stain proteins tagged in this manner. However, we have not tried to detect these tags on a gel directly with the 6xHis Tag Stain.
Will this kit stain the protein in E. coli that has an affinity for the 6xHis tag?
Should this occur, it would appear as a 15K band.