1.Prepare dye reagent by diluting 1 part Dye Reagent Concentrate with 4 parts distilled, deionized (DDI) water. Filter through Whatman #1filter (or equivalent) to remove particulates. This diluted reagent maybe used for approximately 2 weeks when kept at room temperature.
2.Prepare three to five dilutions of a protein standard, which is repre-sentative of the protein solution to be tested. The linear range of theassay for BSA is 0.2 to 0.9 mg/ml, whereas with IgG the linear rangeis 0.2 to 1.5 mg/ml. (See Common Questions, question 4, for moreinformation.)
3.Pipet 100 µl of each standard and sample solution into a clean, drytest tube. Protein solutions are normally assayed in duplicate or tripli-cate.
4.Add 5.0 ml of diluted dye reagent to each tube and vortex.
5.Incubate at room temperature for at least 5 minutes. Absorbance will increase over time; samples should incubate at room temperature forno more than 1 hour.
6.Measure absorbance at 595 nm.