Sensitivity and variability of the Bradford protein assay in the presence of detergents.
The effects of Triton X-100, sodium dodecyl sulfate (SDS), and urea on the response of Coomassie blue G to 16 different proteins and peptides of Mr 1140 to 146,000 were studied to assess the significance of protein conformation and of ionic and nonionic interactions for the dye response to individual proteins. Triton X-100 at a final concentration of 0.008% (v/v) increased the sensitivity of the Bradford assay toward all proteins of Mr 5700 or higher by an average 33%. Increases ranged from +11% with myelin basic protein to +128% with aprotinin. The relative range of absorbance of proteins and deviations from bovine serum albumin decreased by approximately 25%. Triton X-100 appears to facilitate nonionic interactions of the dye with proteins of limited capacity for ionic binding. Conformation of proteins also seemed to be of some significance because the chaotropic agent urea (0.16 M final concentration) increased sensitivity of the assay by 14%. Sensitivity of the assay was lowered by SDS (0.004% final concentration, w/v) by an average 75% from that of the control assay. The results indicate that the incorporation of low concentrations of a nonionic detergent may be useful in improving sensitivity and variability of the Bradford assay.
Friedenauer
S
S
Institute of Pathochemistry and General Neurochemistry, Heidelberg, Federal Republic of Germany.
Berlet
H H
HH
D003902:Detergents; D017830:Octoxynol; D011092:Polyethylene Glycols; D011506:Proteins / Q000032:analysis*; D012394:Rosaniline Dyes; D012967:Sodium Dodecyl Sulfate; D013194:Staining and Labeling; D013501:Surface-Active Agents; D014508:Urea
D003902:Detergents; D011506:Proteins; D012394:Rosaniline Dyes; D013501:Surface-Active Agents; D012967:Sodium Dodecyl Sulfate; D011092:Polyethylene Glycols; D014508:Urea; D017830:Octoxynol; C004692:coomassie Brilliant Blue