Bio-Rad DC™ Protein Assay - Manual (Page 6)

4. Add 25 µl of reagent A' or reagent A (see note from step 1) into each well.
5. Add 200 µl reagent B into each well. If microplate reader has a mixing func-
tion available, place plate in reader and let the plate mix for 5 seconds. If not,
gently agitate the plate to mix the reagents. If bubbles form, pop them with a
clean, dry pipet tip. Be careful to avoid cross-contamination of sample wells.
6. After 15 minutes, absorbances can be read at 750 nm. The absorbances will
be stable for about 1 hour. (See Troubleshooting Guide for recommendation
on using a wavelength other than 750 nm.)
Section 6
Response of Various Proteins
As with any colorimetric assay, different proteins will elicit greater or less-
er color formation. The following proteins have been assayed with the protein
assay. As demonstrated by the graph, there is a slight variation in color devel-
opment with different proteins.
Section 7
Storage
Lyophilized preparations of Protein Standard I (bovine gamma globulin) and
Protein Standard II (bovine serum albumin), if included, should be refrigerated upon
arrival. These lyophilized preparations have a shelf life of one year at 4 °C. Rehydrated
and stored at 4 °C, the protein solutions should be used within 60 days. Rehydrated
and stored at -20 °C, the protein solutions should be used within 6 months.
REAGENT A, REAGENT B, and REAGENT S should be stored away from
direct sunlight at room temperature (25-30 °C). (Reagents A and B may also be
stored in the refrigerator.) All reagents are good for 6 months from date of purchase.
protein concentration, mg/ml
absorbance
BSA
IgG
Ribonuclease A
Ovalbumin
Conalbumin
2.0 1.0 0.0
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