Thermo Scientific Pierce Coomassie Brilliant Blue R-250 and G-250 are purified powders of the two most common forms of coomassie dye, which are key components 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 595nm).
- Easy detection – Develops intensely colored complexes with proteins
- High sensitivity – Can determine as little as 0.5µg/cm2 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 549nm
|Chemical structures of coomassie brilliant blue dyes R-250 and G-250.
|Chemical properties of Coomassie Blue R250:
- Name: Coomassie Brilliant Blue R-250
- Formula: C45 H44 N3 Na O7 S2 (sodium salt)
- CAS number: 6104-59-2
- Molecular weight: 825.97
- Solubility: essentially insoluble in water; dissolve first in methanol
|Chemical properties of Coomassie Blue G250:
- Name: Coomassie Brilliant Blue G-250 (colloidal)
- Formula: C47 H48 N3 Na O7 S2 (sodium salt)
- CAS number: 6104-58-1
- Molecular weight: 854.02
- Solubility: slightly soluble in water; best to dissolve first in methanol or ethanol
Recipe for simple coomassie protein stain.
45% Methanol (reagent grade)
10% Glacial acetic acid
3g/L Coomassie Brilliant Blue R250
- Add 100mL of glacial acetic acid to 450mL ultrapure water.
- Dissolve the 3g of Coomassie Dye in 450mL methanol.
- Filter the solution before use.
Syrovy, L. and Hodny, Z. (1991). Staining and quantification of proteins separated by polyacrylamide gel electrophoresis. J. Chromatog. 569, 175-196.
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