Bio-Rad ReadyPrep Sequential Extraction Kit - Manual (Page 14)

Modified Bio-Rad Protein Assay
This modification to the standard Bio-Rad protein assay
procedure (catalog # 500-0001) is recommended to deter-
mine the protein content of Extracts 2 and 3 (modified from
[10] and [11]). It is not necessary with Extracts 1, but it can be
used with Extracts 1, the content of the samples, not the pro-
cedures. The high concentrations of detergents and
chaotropes in Reagents 2 and 3 interfere with many other
types of assays.
1 Prepare bovine gamma globulin (BgG) standards at about
14 mg/ml.
• Rehydrate protein standard I with 2 ml of deionized water.
Disregard the instructions with the standard that call
for rehydrating this protein with 20 ml of water. The
protein concentration of standard, when rehydrated
with 2 ml of water, will be 10 times the concentration
shown on the bottle.
2 Prepare a 1:4 dilution of dye reagent concentrate.
• Mix 1 part of the dye reagent with 3 parts of water and
filter the diluted dye through Whatman Number 1 filter
paper, or equivalent.
3.5 ml of diluted dye reagent are used in each standard
and assay tube. Prepare excess dye.
3
Prepare 0.12 N HCl (nominal) by diluting concentrated HCl
1:100 with water.
• Add 10 µl of concentrated HCl to 1 ml of water.
•
80 µl are used in each assay tube. Prepare excess.
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