Bio-Rad Protein Assay - Manual (Page 7)

5
Fig. 1. Typical standard curve for the Bio-Rad Protein Assay, bovine gamma
globulin (standard I), bovine serum albumin (standard II).
O.D.
595
corrected for
blank - 200-1,400 µ g/ml x 0.1 ml = 20-140 µ g protein.
2.3 Microassay Procedure
1. Prepare three to five dilutions of a protein standard which is represen-
tative of the protein solution to be tested. The linear range of the assay
for BSA is 1.2 to 10.0 µg/ml, whereas with IgG the linear range is 1.2
to 25 µg/ml. (See Common Questions, question 4, for more informa-
tion.)
2. Pipet 800 µl of each standard and sample solution into a clean, dry
test tube. Protein solutions are normally assayed in duplicate or tripli-
cate.
3. Add 200 µl of dye reagent concentrate to each tube and vortex.
4. Incubate at room temperature for at least 5 minutes. Absorbance will
increase over time; samples should incubate at room temperature for
no more than 1 hour.
5. Measure absorbance at 595 nm.
Standard procedure
(20-140
µ g)
Protein (mg/ml)
BSA
IgG
O.D
. 595
0.20
0.2
0.4
0.6
0.40 0.60 0.80 1.00 1.20 1.40
0.8
1.0
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