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Boster’s Commassie Blue Staining And Destaining Solution is used for staining and destaining of protein on polyacrylamide gels.
|Pack size||1kit (for 10 assays to stain gels of 5 x 8.5cm)|
|Applications||For protein staining on polyacrylamide gels.|
|Commassie Blue Staining Solution||1||100mL||0.1% (w/v) Brilliant Blue R-250, 25% (v/v) methanol and 5% (v/v) acetic acid||AR0163-1|
|Commassie Blue Destaining Solution||1||500mL||25% (v/v) methanol and 5% (v/v) acetic acid||AR0163-2|
|Product Name||Commassie Blue Staining And Destaining Solution|
|Size||1kit (for 10 assays to stain gels of 5 x 8.5cm)|
|Working Concentration||10mL for each gel|
|Storage||Upon receipt store at room temperature. It is stable at room temperature for one year.|
|Label or Dye||Coomassie Blue R-250|
|Equivalent||Millipore Sigma (Product No. B8522)|
|Application||For protein staining on polyacrylamide gels.|
|Compatibility||Compatible with mass spectrometric analysis|
|Cite This Product||Commassie Blue Staining And Destaining Solution (Boster Biological Technology, Pleasanton CA, USA, Catalog # AR0163)|
Usually after electrophoresis, researchers should evaluate the result through staining and destaining, and detect the protein which was disassembled into different bands from the gel. Besides, researchers also could electroblotting the gel for different research purpose. This solution is a coomassie blue R-250 dye-based, can be widely used for staining of protein electrophoresis gel of SDS-PAGE or non-denaturing PAGE, as well as in the residual protein detection of PAGE gel after by Western transferring. By this means, the working range for protein electrophoresis bands can be as low as 10ng. This production is acidic and its corrosion requires a necessary protection when use it.
• Easy detection—Develops intensely colored complexes with proteins
• High sensitivity—Can determine as little as 10 ng of protein present in a gel matrix
• Reversible staining—Anion of Coomassie Brilliant Blue formed in the acidic staining medium combines with the protonated amino groups of proteins by electrostatic interaction; resulting complex is reversible under the proper conditions
• Differentiation between bound and unbound dye—When dissolved in 0.01M citrate buffer at pH 3.0, has an absorption maximum at 555nm; protein-dye complex is characterized by a peak slightly broader than that of the free dye with a maximum at 549 nm
Coomassie Brilliant Blue R-250 is one of the most common forms of coomassie dye, which is a key component of various colorimetric protein gel stains. Coomassie R-250 and G-250 dyes are two chemical forms of a disulfonated triphenylmethane compound that is commonly used as the basis of stains for detection of proteins in gel electrophoresis and Bradford-type assay reagents for protein quantitation. The R-250 (red-tinted) form lacks two methyl groups that are present in the G-250 (green-tinted) form, which is also called colloidal coomassie dye. Typically, coomassie gel stains and protein assay reagents are formulated as very acidic solutions in 25 to 50% methanol. In acidic conditions, the dye binds to proteins primarily through basic amino acids (primarily arginine, lysine and histidine), and the number of coomassie dye ligands bound to each protein molecule is approximately proportional to the number of positive charges found on the protein. Protein-binding causes the dye to change from reddish-brown to bright blue (absorption maximum equals 595 nm).
1. After electrophoresis, place gel in a clean tray. Wash the gel with 100mL of distilled water 3 times for 5-10 min each with gentle shaking. This step is to gently remove the interference of SDS and salt which interfere with binding of the dye to the protein.
2. Discard last wash. Add sufficient volume of Commassie Blue Staining Solution to cover the gel.
Note: Shake the Commassie Blue Staining Solution before use.
3. Incubate at room temperature for 60 minutes with gentle shaking. When the color of the gel is very close to that of the staining solution, the gel is stained sufficiently.
Note: To speed up staining, heat the staining container in a microwave to near boiling (try to avoid boiling to prevent gel breaking), immediately stop heating, place on a shaker and shake for a few minutes.
4. Pour out the staining solution and wash the gel with distilled water.
Note: The staining solution can be reused for 2-3 times.
Note: For the gel thicker than 1.5mm, it takes a longer period for above mentioned washing and staining steps.
5. Add Commassie Blue Destaining Solution and incubate with gentle shaking until background is clear. Change Commassie Blue Destaining Solution during incubation.
Note: To speed up destaining, heat the staining container in a microwave to near boiling (try to avoid boiling to prevent gel breaking), immediately stop heating, place on a shaker and shake for a few minutes.
6. Store the gel in water containing 20% glycerol.
|No band is seen||Sample is not enough||Try to add two BSA lanes with different volumes when electrophoresis as a positive control.|
|High background||Impurity is not removed completely||Try to wash longer or wash more times|
|Precipitate occurs when staining the gel||the container was polluted||Try to use a clean container while using new stain solution until the bands are seen.|
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